West Coast 6th PLANT BREEDING SYMPOSIUM CONTENTS CLUB MYKONOS Resort

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					                    6th PLANT BREEDING SYMPOSIUM


                                    CONTENTS



CLUB MYKONOS: Resort information & conditions                    1

WORKSHOP
    Programme                                                    2
    Introduction to I.P. systems                                 3
            Plant breeders’ rights                               5
            New developments                                     10
    Patent rights                                                12
    Trade mark and geographical names                            13
    Biodiversity and traditional knowledge                       15
    Options for shared I.P. in partnership/consortium research   16
    Case studies                                                 18

SYMPOSIUM
    Programme                                                    22
    Posters                                                      30

ABSTRACTS
     Keynotes                                                    32
     Oral Presentations                                          41
     Poster Presentations                                        94

DELEGATE LIST                                                    112
                                                                                                             1




West Coast                                                            Tel: 022 7077000
Holiday Lifestyles (Pty)ltd                                           Fax: 022 7077783
Private Bag x 2                                                       VAT. No: 4180203327
LANGEBAAN                                                             Reg. No: 1994/0713/07
7357                                                                  conferences@clubmykonos.co.za



                              RESORT INFORMATION & CONDITIONS


The right of admission to the Resort is reserved at all times.

Kalivas are fully equipped. Own toiletries to be provided

Kalivas are allocated according to availability. Requests in relation to specific location (i.e. Sea
facing) cannot be guaranteed.

Kalivas can only be occupied from 16h00 and must be vacated (i.e. key handed in at reception) at
10h00 on the day of departure.
A late departure fee of R100.00 will be levied.

Portage service is not provided

Beach towels are not provided.

All leisure facilities are available for residents, but please note that some activities do have a cost to
use.
Bathing caps are compulsory for swimming pools.

Keys are available at Reception on arrival. Hair dryers, safe keys and irons are also available at
Reception at a R50 deposit fee per item, and TV Remotes at R200. (LIMITED TV REMOTES)

Pets are not permitted on the Resort

Kalivas are serviced daily, except Sundays and Public Holidays.

All bar and restaurant services are direct cash or credit card only. No signing to your account
allowed

PARKING AT OWNERS RISK

Any enquiries – contact ext 9 (Swithboard)
                                                                                      2


                            PROGRAMME - WORKSHOP

                                MONDAY 13 MARCH 2006
10:00-11:00    Registration

11:00          Workshop – Genetic resources and your intellectual property
               Presenters: Bastiaan Koster, Patent lawyer, BowmanGilfillan
                           Wynand van der Walt, Senior partner, FoodNCropBio

11:00-11:25    Introduction to I. P. systems (B. Koster)
               (covers systems, benefits, WIPO, WTO-TRIPS)

11:25-11:50    Plant breeders’ rights (W. van der Walt)
               (covers UPOV, farm-saved seed, essential derivation, PBR status in RSA,
               enforcement, how to apply)

11:50-12:10    New developments (W. van der Walt)
               (covers SADC Model Law on PBR, South African IPR policies, Upgrading
               PBR Act)

12:10-12:35    Patent rights (B. Koster)
               (covers requirements, patenting genes, protection, how to apply)

12:35-13:00    Trade mark and geographical names (B. Koster)
               (covers requirements, protection, how to apply)

13:00-14:00 Lunch

14:00-14:30    Biodiversity and traditional knowledge (B. Koster)
               (covers briefly Convention on Biological Diversity, Biodiversity Act, and
               traditional knowledge recognition)

14:30-15:00    Options for shared intellectual property in partnership/consortium
               research (B. Koster & W. van der Walt)

15:00-16:30    Case studies (an inter-active session) (W. van der Walt & B. Koster)
          a.   Plant Breeders’ Rights—Patents—Trade marks interaction
          b.   Modern biotech complications—What is patentable?
          c.   Rooibos trade mark
          d.   Sugraone grapes
          e.   Protection and infringement in fruit trees
          f.   Policing infringements on seed varieties
          g.   Breeding from a F1 hybrid
          h.   Breeding with indigenous plants
          i.   Barley cv. Franklin: Cultivaust vs Tasmania
          j.   Other cases and questions from participants

16:30:         Closure
                                                                                           3


          INTRODUCTION TO INTELLECTUAL PROPERTY SYSTEMS
                                     Bastiaan Koster

Philosophy behind the existence of registered IP rights and the building blocks thereof:


•    Patents

•    Trade Marks

•    Plant Breeders Rights

•    Registered Designs

•    Copyright

•    Know-how and Trade Secrets

•    Unlawful Competition and Passing-off.


Balancing of IP rights
How does the world propose to deal with conflicting concepts of “monopolies” on the one
hand, against the need for “free competition” on the other. The trends and latest
developments


WORLD INTELLECTUAL PROPERTY ORGANISATION (WIPO)


The function of WIPO
Developing countries have made demands through WIPO for changes to IP systems which
will better serve the needs of developing countries. There is a perception that IP systems
have only benefited developed countries.


Development Agenda at WIPO
Developing countries have blocked harmonisation efforts of WIPO on the patent system.


Possible Harmonisation between the USA, European and Japanese Patent offices
                                                                                             4


The so-called “World Patent” and other developments to simplify and reduce the costs
of registration of IP rights
   •   Europe – the Community Patent – probably at least eight years from being finalised.
   •   Trade Marks – treaties like Madrid and the Community Trade Mark system makes it
       cheaper and easier to obtain registered trade marks in a number of countries.
   •   For the foreseeable future, IP rights will remain territorial and only in some instances
       regional.
                                                                                                5


                              PLANT BREEDERS’ RIGHTS
                                    Wynand van Jaarsveld

Article 27 of the Universal Declaration of Human Rights:
“… it guarantees everyone’s right to the protection of moral and material interests
resulting from any scientific, literary or artistic production of which he/she is the
author”


The Union For the Protection of New Plant Varieties was established in Paris in 1961, came
into force in 1968, and was revised in 1972, 1978, and 1991. This Convention established a
global regime for plant breeders’ rights for varieties that are new, distinct, uniform, stable and
have an acceptable designation. Its mission is to manage a global harmonized system that will
promote the development of new plant varieties for the benefit of society.


Under the UPOV Convention, protection of plant varieties is granted if the variety is new, i.e.
not known in the market for over 1 year in country developed or over 4 years (6 for
trees/fruit) in other convention countries, distinguishable in terms of botanical description
from another known varieties, sufficiently uniform in appearance, stable over cycles of
propagation or multiplication, and have an acceptable denomination (name). Other salient
requirements are
   •   All species on official lists are eligible (extended in 1991 to all plant species)
   •   Applications are made per country or region
   •   The government shall examine the documentation and may grow out plants for
       examination
   •   Protection covers production, offering for sale, marketing, imports, exports, of the
       propagating material. UPOV 1991 extends protection to harvested products of the
       variety in cases of unauthorized use of the propagating material.
   •   The protection period shall be at least 15 years for seed crops and 20 years for tree
       and vine species (extended to 20 and 25 years under 1991)
   •   The rights of the breeder shall not extend to further breeding by others
   •   A member state of the Convention must extend same rights to other member states
   •   NOTE: Distinctness is based on phenotype
                                                                                               6




The 1991 revision of UPOV also clarified and/or added the following aspects:
   •   Article 14.5 extends protection to essentially derived varieties i.e. new varieties
       derived from, but distinguishable from, initial protected varieties, when the derived
       variety retains the essential characteristics (i.e. genetics) of the initial variety.
   •   NOTE: EDV is based on genome i.e. looks different but has essentially retained
       genome
   •   Article 15.1 provides for compulsory exceptions from protection as acts done for
       private and non-commercial purposes, experimental acts, and further breeding
       (subject to Article 14.5).
   •   Article 15.2 defines that within reasonable limits, subject to safeguarding the
       legitimate interests of the breeder, a farmer may use for propagating purposes on their
       own holdings, harvested material obtained from a protected variety planted on their
       own holding. This exemption does not entitle the farmer to sell seed. The problem is
       that neither reasonable limits nor legitimate interests are defined and compliance is
       left to national governments.. The US permits farm-saved seed, while most EU
       countries and Australia are trying to mandate payments of royalties on such farm
       seed. The EU has published extensive regulations that require royalties of between 40
       and 50% of standard royalty rates to be paid on farm-saved material, exempting
       small-holder farmers according to a formula of volume of crop produced.


In 2005 UPOV developed a Model Law that provides more flexibility for developing
countries to become members. Membership stood at 60 states at September 2005, including
the EU as a block (22 states), bringing the total number with UPOV or UPOV type PBR at
over 100. On January 1, 2006, the West African group of 16 states under the African
Intellectual Property Organization became full members.


NOTE: The US Plant Variety Protection Act of 1970, updated in 1994 to meet UPOV 91,
applies to sexually propagated plants, hybrids, and tuber propagated varieties. The US Patents
Act provides patent protection on asexually propagated plant varieties.
                                                                                              7


South Africa drafted a PBR Act in the mid sixties and updated it in 1976(Act 15) to meet
UPOV 1978 to become the 10th UPOV member in 1978. The Act was amended in 1996 to
meet UPOV 1991 but South Africa never acceded to the 91 Convention. Article 23A was
poorly written so that it seems that no protection exists in the first cycle of farm-saved
material. Steps in applying for PBR are as follows:
     •   Application by citizen or authorized licensee/agent
     •   Variety on list of eligible species
     •   Acceptable denomination
     •   Completion of application form and technical questionnaire
     •   Accompanied by payment of fees
     •   Sample of plant propagating material to be submitted
     •   PBR and variety listing can be done same time
     •   NOTE: Applications and decisions are published, objections are possible, once
         granted PBR remain in force for term subject to payment of annual fees.
     •   Fees are stipulated for 18 types of applications, including R1000 for an application
         for PBR, R1700 to R2200 fee for examination, and annual fee of R200


Strengthening of PBR can make use of compulsory certification, licensing/contacts, growers
clubs, and employing an enforcement agent. Patents and Trade Marks can be used, subject to
eligibility.


Key parameters of the South African PBR status as at December 2004 are as follows (this
information has FANRPAN copyright):
               •   The 1807 PBRs per crop group are 236 for 17 species of vegetables; 774 for 67
                   ornamental/flower species; 244 for 28 fruit species; 442 for 24 agronomic
                   (grain, oil, protein, industrial crops) species; and 111 for 23 forage/grass
                   species.
               •   The species with the highest PBR count is roses with 356 varieties
               •   The data base of the Registrar of PBR shows a total number of 1133 parties
                   that have been applying for PBR. Most of these are small operators which
                                                                                    8


    proves that South African PBR is not dominated by a few large local or a few
    multinational companies
•   South African breeders own a total of 704 varieties; 344 by private companies
    and breeders, 87 by multinational subsidiaries based in South Africa, 259 by
    the Agricultural Research Council, 10 by universities, and 4 by one
    biodiversity institute. This shows the benefit of PBR for successful investment
    in breeding by local private breeders
•   South African ownership of PBR varieties comprises 39 per cent of the 1807
    total. Private South African varieties are 431 and public varieties are 237,
    showing the investment by the private sector in plant breeding
•   Analysis of listed fruit varieties shows that 34 varieties had PBR out of 89
    apple listed as scion material; 2 had PBR out of 11 rootstock varieties; 6 out of
    16 apricots had PBR; 1 out 5 prune varieties; 21 out of 48 nectarine; 37 out of
    118 peach; 23 out of 47 japanese plum; all 3 plumcots; 5 out of 18 Prunus cross
    rootstock;13 out of 42 pear; 2 out of pear rootstock; 32 out of 186 table grape;
    and 5 out of 44 table grape rootstock varieties had protection under PBR.
•   Insight into stimulus for breeding varieties of open-pollinated, self-pollinated
    and vegetatively propagated crops was obtained by comparing PBRs granted
    over 15 years for a few select crops where profit margins are low and farm-
    saved seed is common. PBRs granted over 4 years from 2001-2004 were
    compared against total over 15 years since 1990. For wheat the data show that
    26 (45 %) of the total PBRs granted since 1990, were granted in the past 4
    years; for triticale the figures were 5 varieties (42%) in the last 4 years; for
    potatoes 25 (41%) in the last 4 years; for dry beans 20 (43%) in the last 4 years;
    and for soyabeans 44% in the last 4 years. Therefore, companies continue to
    breed new varieties although their persistence may be limited
•   Comparison of the last 5 years of fruit PBRs granted with the past 20 years
    yielded a different picture. For citrus the figure was 55 %, apple 22%, peach
    9%; nectarine 4%, and table grapes 36% of the total registered in the last 5
    years
                                                                                     9


•   Plant breeders, in general, do not see much value in registering PBR on hybrids
    as the protection lies in confidential information on the pedigree i.e.
    combination of parent lines. Farm saved seed will result in segregation of
    beneficial traits and loss of hybrid vigour. Yet, following PBR on new open-
    pollinated varieties that started in 1992, some 74 PBRs (88% of the 84 total)
    were registered in the last 5 years, mostly for hybrids. The reasoning seems to
    be that only a minimal fee is charged when application and testing for
    distinctness, uniformity and stability take place simultaneously for official
    variety listing and PBR. Thus, some protection is gained for minimal expense.
    Breeders have not yet applied for PBR on inbred lines due to risk on
    compulsory licensing and risk of inbreds having to be planted out in
    government tests, as well as the risk of applications for compulsory licences.
                                                                                           10


                                NEW DEVELOPMENTS
                                   Wynand van Jaarsveld

WTO-TRIPS
WTO has a mission to facilitate global trade by reducing trade barriers. Article 27 of the
Trade Related Intellectual Property Systems Agreement requires that “patents shall be
available for any inventions, whether products or processes, in all fields of technology,
provided that they are new, involve an innovative step, and are capable of industrial
application. Paragraph 27.3.b states that: “members shall provide for the protection of plant
varieties either by patents or by an effective sui generis system, or by a combination
thereof”. The same paragraph also provides that members may exclude for patent eligibility
“plants and animals other than micro-organisms, and essentially biological processes for
the production of plants and animals, other than non-biological and micro-biological
processes”. It is in practice only the US (and by exception Australia and Japan) that award
patent rights on plant varieties. The sui generis option means a protection system “of its own
kind” which provides a flexible approach.
       A great many WTO members, especially in Africa, are now scrambling to get their IP
systems in place.


SOUTH AFRICA
All IPR legislation is being revisited by government in order to create more harmony, to
make provision for protection of indigenous/traditional knowledge and farmer varieties/land
races, and to enable previous disadvantaged sectors of the population to become part of IP
systems and derive due benefits. The Department of Agriculture has an advanced draft policy
on agricultural IPR that is expected to be published soon for comment. It will lead to
amendments in the PBR Act. This opens the way for private, academic and public plant
breeders to be actively involved and to insert provisions that will strengthen PBR.
       The ARC has commissioned a private law firm to develop an IPR policy for them.
The CSIR is working on an improved IP policy, as are many universities. Some universities
are well advanced in shared IP to the point of start-up companies to commercialize their
                                                                                         11


innovations. It is recommended that all plant and animal breeders get themselves involved in
these developments and not be left behind.


AFRICA AND SADC
Only South Africa, Kenya and Tunisia are UPOV members, while 16 West African countries
joined UPOV on 1 January 2006 under the umbrella of OAPI. Zimbabwe has had PBR since
the late 1960s but did not accede to UPOV. Their later application required some
amendments in order to meet UPOV 1978. In contrast, almost all Latin American countries
are UPOV members. UPOV has drafted a Model Law specifically to facilitate entry by
developing countries. One key provision in this model, as in the OAPI model under the
Accord de Bangui, is that new entry countries will have one year during which existing
known/commercial varieties can be placed on the PBR list, subject to meeting distinctness
requirements.
       SADC is slowly awakening but PBR development is very slow and often
contaminated by blending in other issues such as seed certification, variety listing,
commercial value for agriculture, and local seed production. The issues of indigenous
knowledge and customs, and farmer‘s privilege/rights rest on different legal principles and
combination with PBR often lead to further weakening protection for breeders. (as in the
African Union Model Law).


The SADC status on PBR is as follows:
      No PBR: Botswana, Lesotho, Namibia, Swaziland
      First draft: Angola, DRC, Mozambique, Zambia
      Advanced/final draft: Malawi, Mauritius, Tanzania
      UPOV application: Zimbabwe
      UPOV member: South Africa

An initiative has been launched by FANRPAN, the regional agricultural policy analysis and
development organization to study the present status of IPR in SADC, and to develop
proposals. One outcome was a draft model law on PBR for the region and to strive for a
regional block to join UPOV, envisaging a regional office for PBR.
Again, all plant breeders are encouraged to become involved in this process at national and
regional level.
                                                                                           12


                                      PATENT RIGHTS
                                      Bastiaan Koster


Invention must be new, inventive and capable of being used in trade industry or
agriculture to be patentable


Novelty requirements – what does it mean?


Inventiveness – what does it mean?


Section 25(4) provides that “a patent shall not be granted;


For any variety of animal or plant or any essentially biological process for the production of
animal or plants, not being a micro biological process or the product of such a process.


Micro-biological processes and their products relating to animals or plants are
therefore patentable
What is a micro-biological process?


Obtaining a patent and a plant breeder’s right for a genetically modified plant. Is this
possible?


The patenting of genes – what are the developments in Europe?
                                                                                       13


                   TRADE MARK AND GEOGRAPHICAL NAMES
                                      Bastiaan Koster


The purpose of a trade mark is to distinguish the products or services of the owner of the
trade mark from similar products or services belonging to another.


A trade mark must therefore be capable of DISTINGUISHING.


A trade mark can be inherently capable of distinguishing or it can become capable of
distinguishing for use.


As a general rule, descriptive names are not capable of distinguishing and therefore not
registrable as trade marks. Descriptive type trade marks may however become capable of
distinguishing through use. For example, “the Hi Fi Corporation”.


Brand Equity – What does it mean and why is it important?
What are the considerations in selecting a trade mark for a new product or service?


•     Why is the trade mark needed?

•     What are your competitors doing?

•     What other good / strong trade marks exist in that particular area?

•     Is it a consumer product or service or only sold to a selected clientele?

•     Looking at other similar trade marks

•     Must have a strategy



Trade Mark issues in the plant and food industry
                                                                                             14


GEOGRAPHICAL NAMES


Appellation of Original
Refers to a sign that indicates that a product originates in a specific region but is limited to
those cases where the characteristic qualities of the products are due to the geographical
environment, including natural and human factors in that region, for example, “Roqueford”,
“Gorgonzola” or “Habana”.


Geographical Indication
A term that is used broadly for all forms of protection for indications of geographical origin.
It is, however, more than a mere indication of source.


What is the relevance?


Never use the geographical name as a trade mark.


•     The South African and European Union Agreement
                                                                                          15


               BIO-DIVERSITY AND TRADITIONAL KNOWLEDGE
                                     Bastiaan Koster

Convention on Biological Diversity (CBD)
The CBD has been signed by some 170 countries with the notable exception of the United
States of America.


The objectives of the CBD are stated broadly as the following:


•    The conservation of biological diversity;

•    The sustainable use of its components and the fair and equitable sharing of the benefits
     arising out of the utilisation of genetic resources, including by appropriate access to
     genetic resources; and

•    Appropriate transfer of relevant technologies, taking into account all rights over those
     resources and to technologies and by appropriate funding

Update on the South Africa Bio-Diversity Act




Traditional Knowledge

•    Update on discussions at WIPO relating to Traditional Knowledge, Genetical
     Resources and Traditional Cultural Expressions. Where is all of this going?
                                                                                              16


  OPTIONS FOR SHARED I.P. IN PARTNERSHIP/CONSORTIUM RESEARCH
                            Bastiaan Koster/Wynand van der Walt


Most countries have realized that they do not have enough critical mass to do research and
derive benefits in a global environment dominated by multinationals from industrial
countries. Yet, many developing countries have a core fraternity of dedicated scientists who
do world class innovation. There is a strong trend towards consortium projects and private-
public partnerships, at national, regional and international level. A negative trend is declining
government funding for agricultural research. The old policy that all IPR rests with the public
institute or university, irrespective of where the funds come from, has no room in the new
millennium. No partnership can work on this basis.


One excellent example is the research/innovation/IPR policies of the University of Wisconsin
in the US, now entering its 78th year of successful partnerships. The University set up WARF
(the Wisconsin Alumni Research Foundation) with the objective of getting their innovation
and technologies into the market by way of the private sector. This was followed in
subsequent years by a revenue producing industrial relations unit, a technology transfer
office, a funded incubator for 50 start-up companies, and a research park that employs some
3500 people housed in 34 buildings (bigger than our CSIR). There are several surveys in the
US that show win-win situations for all, amounting to billions of dollars.


The US Bayh-Dole Act of 1980 paved the way for universities to take out IPR on their
innovations and products, and to enter into licence agreements. A 1999 survey showed that
190 institutions had filed 8800 patent applications. Gross income from licensing was $862
million end economic impact $41 billion, while 271 000 jobs were created.


It is up to scientists to convey the message to policy makers that IPR based on innovation is a
major driver of the economy. South Africa seems to be in its infancy when it comes to
deriving benefits from IPR in agriculture.


Like any good business arrangement, it should be practical, be capable of execution and fair
on the parties concerned.
                                                                                         17


Various options will be discussed.

Considerations to be taken into account:

   •   Intellectual contribution by a party
   •   Financial contribution by a party
   •   Length of time of the research. How quickly can the research be commercialised.
   •   Risks undertaken by parties. Barriers to entry by others
   •   Scope of intellectual property rights.
                                                                                           18


                                     CASE STUDIES
                           Bastiaan Koster/Wynand van der Walt


CASE STUDY 1 - Modern biotechnology complications


Most modern biotech varieties enjoy IP protection under Patents, Trade Marks and PBR. The
South African Patents Act 38 of 1997 follows that of most other industrial countries (US
excepted, and Japan and Australia) in that plant and animal varieties cannot be patented.
Chapter V (B) reads that excluded from eligibility is “any animal or plant variety or any
essentially biological process for the production of an animal or plant, not being a micro-
biological process or product of such a process”.


Do you understand what an essentially biological process is?
What are your opportunities and constraints in breeding from such a GM modern variety?
Can you patent a novel plant cell culture?


CASE STUDY 2 - Rooibos tea trade mark


CASE STUDY 3 - Sugraone grape


CASE STUDY 4 - Breeding from an F1 hybrid
New homozygous inbred lines can be developed by selfing and selecting from hybrids,
populations, open-pollinated varieties, or cross-breeding/backcrossing with existing inbreds.
Breeders need to understand constraints imposed by intellectual property rights and legal
uncertainties from using genetic material owned by private companies (or institutions that
claim ownership). Taking out PBR on hybrids is not common as the owner has the
constitution of the hybrid (pedigree) as a company secret, neither is PBR on parental inbred
lines although the latter may be gaining in importance, since the inbreds are not sold and are
not in the public commercial domain.
                                                                                               19


EXAMPLE: An F1 maize hybrid owned by a private company SeedX , is a single cross (A x
B) of which the two inbreds are SeedX proprietary genetic material. The gene construct for
unique insect resistance is protected by a patent, and this unique trait is further protected by a
trade mark “Wormresist” ®. The hybrid is protected by PBR and it is registered on the
official national variety list. The breeder decided to set up his own private breeding
programme, having been employed previously by SeedX, Consider the following scenarios in
terms of ethics, intellectual property protection under UPOV, patents, and trade marks. There
is no restraining agreement from SeedX to prevent the breeder from engaging in maize
breeding for commercial purposes.


Which of the following actions represent infringements?
   1. Obtained seed from SeedX breeding nursery or having taken seeds of inbred lines
       when he left the employ of the company.
   2. Arranged with contracted grower for SeedX foundation seed production (inbred
       multiplication) to buy as grain for feed purposes the portion of seed that was screened
       out as small and damaged kernels.
   3. Passed by the SeedX’s contracted hybrid seed production field and having recognized
       the inbred lines, took some ears from the pollinator rows.
   4. Bought several bags of commercial hybrid seed from an authorized dealer and planted
       out several hectares. Identifying and selfing weak plants that may be seed parent
       inbred that arose from unplanned self-pollination during the production, or pollinator
       inbred that was due to unintentional mixing when the seed parent rows (now being
       hybrid seed) were harvested. After crossing retrieved inbreds with another inbred,
       obtained several good recovered, slightly modified inbreds through several cycles of
       inbreeding and selection, and proceeded to make hybrids that are minor variants of
       SeedX commercial hybrids.
   5. Selfing hybrid plants of which the seed had been obtained by purchasing hybrid seed,
       and through several cycles of inbreeding and selection, developed new inbred lines.
   6. Being experienced with SeedX hybrid pedigrees and their inbred lines, used pollen to
       obtain hundreds of di-haploids through regenerating haploids and doubling
       chromosome number, selected towards phenotype of known SeedX inbreds. This
                                                                                            20


        process was assisted by genetic fingerprinting and marker-assisted breeding to capture
        major genes originally present in SeedX inbreds.
    7. Being able to guess the parental inbreds used in hybrids by looking at the hybrid
        phenotypes of new commercial hybrids released after he left SeedX, the breeder
        proceeded to cross the hybrids with closely related inbreds in the public domain, and
        selected variants of the SeedX parental inbreds.
    8. Buying seed from several seed companies, knowing that many modern hybrids are
        pure or modified single crosses, making four-way hybrids in crossing blocks and
        using and/or selling such seed.


CASE STUDY 5 - EDV dispute in Gypsophila
Party X has a variety Dan that is protected under the EU PBR registration.
Party Z has obtained PBR under the EU system for two varieties, Blanc and Suns, and is
marketing both.
Party X lodged litigation claiming that Blanc and Suns were essentially derived from his
variety Dan. He produced DNA fingerprint data to support his claim.
Party Z supported his defense by submitting DNA fingerprinting data that differed from those
of Party X. He also claimed that his variety Blanc was found by the Registrar to differ in 17
characteristics from Dan. In the case of Suns, the Registrar did not consider it necessary to
include Dan as comparator.
What do you think the judge should take into account?


CASE STUDY 6 - Sale of barley variety “optic”
Company X had an exclusive licence from a UK breeder for producing and selling this PBR
protected barley variety in NZ He sued company Y and its two directors for “arranging the
sale” of Optic seed, without having a sub-licence or paying royalties. The defendants claimed
that they did not sell the seed. The claimant said that seed does not sell itself. The seed was
“dispensed of” in plain bags, some as feed and others for sowing. There was no clear paper
trail of the transactions.
You be the judge! What grounds would you use for a decision?
                                                                                            21


CASE STUDY 7 - Sale of PBR protected barley seed
Company A had an exclusive licence from the Dept Agric in Tasmania to produce and
market their protected barley variety, Franklin. Company A became aware that farmers were
retaining harvested material for use as planting seed but did not take legal action against the
farmers. It was later found that a major co-op was also marketing the variety from commodity
grain. After some time Company A visited the co-op and demanded royalties but took no
immediate legal action when the co-op refused. They subsequently took the co-op to court.
The co-op said that Company A did not arrange for a contract with them to sell seed and to
pay royalties.
What verdict would you as judge make and on what grounds?
                                                                                        22


                    6th PLANT BREEDING SYMPOSIUM

                                   PROGRAMME

                             MONDAY 13 MARCH 2006
10:00-11:00   Registration

11:00         Workshop – Genetic resources and your intellectual property

17:00-17:45   Registration

18:00         Sundowners boat trip (subject to weather)

20:00         Meet and Greet Dinner at Marina terrace


                             TUESDAY 14 MARCH 2006

07:00-08:00   Registration

                                 Session 1 Opening
                             Chair: Dr. Francois Koekemoer

8:15-8:30            Introduction & Welcoming address
                     Dr. Francois Koekemoer: President: Southern African Plant Breeders’
                     Association

8:30-9:00     K1     Monsanto – Invited speaker
                     Breeding for performance under drought; from art to science
                     Maarten van Ginkel
                     Department of Primary Industries, Victoria, Australia; Molecular Plant
                     Breeding Cooperative Research Center


                              Session 2 Abiotic stresses
                                Chair: Mr Antony Jarvie

9:00-9:30     K2     Water quality of the lower Vaal River catchment and its influence on
                     soils and crops
                      Chris du Preez, M.G. Strydom, P.A.L. le Roux, J.P. Pretorius, L.D.
                      van Rensburg and A.T.P. Bennie
                      Department of Soil, Crop and Climate Sciences, University of the Free
                      State
                                                                                           23



9:30-9:45     1.    The influence of nitrogen on polymeric and monomeric proteins and
                    quality in hard and soft wheat
                    M.T. Labuschagne, G. Meintjes and F.P.C. Groenewald

9:45-10:00    2.    Breeding new stone fruit rootstocks with specific stress resistance traits
                    H. Ham

10:00-10:30   Tea/Coffee

10:30-11:00   K3    Climate change and possible effects on agriculture in South Africa
                    Johan van den Berg
                    Business Development: ARS

11:00-11:15   3.    Investigating drought tolerant Eucalypt species and provenances for
                    Northern, Coastal Zululand
                    R. A.W. Gardner and T. Swain

11:15-11:30   4.    Adapting ARC fruit breeding goals to address changes in climate
                    J.P. Human, H. Ham, W.M. Pieterse and I.F. Labuschagnè

11:30-11:45   5.    Estimates of heterosis and association of genetic distance with
                    heterosis in durum wheat under different moisture regimes.K.F.
                    Solomon, Labuschagne, M.T. and C.D. Viljoen

                           Session 3A Breeding techniques
                               Chair: Dr. Brooks Coetzee

11:45-12:00   6.    Some aspects of experimental design in a 10-year old Pinus patula
                    progeny trial
                    A. Kanzler

12:00-12:15   7.    Recurrent mass selection as a means to pyramid resistance genes in
                    wheat
                    G.F. Marais, W.C. Botes, H.S. Roux, J.E. Snyman and M.M.
                    Zaayman.

12:15-12:30   8.    Mechanisation and computerisation of yield testing in a maize
                    breeding programme.
                    R. W. Dunlop and J. D. Rossouw

12:30-12:45   9.    Where eucalypt tree breeding and biotechnology meet - an unfortunate
                    case study!
                    T. Swain

13:00-14:00 Lunch
                                                                                         24



14:00-14:15   10.   Computer modelling: a tool for comparing selection strategies in a
                    simulated poultry population and how this can be used in forestry
                    J. De Guisti, A. Fossey and C. Hancock

14:15-14:30   11.   Update on the multiple population breeding strategy implemented for
                    Acacia mearnsii (black wattle) in South Africa - three-year growth
                    measurements
                    S.L. Beck, J de Guisti, R. W. Dunlop and M. D. V. Resende

14:30-14:45   12.   Developing a non-destructive analytical technique for pulp yield in
                    Black Wattle (Acacia mearnsii) - Part 2
                    R. W. Dunlop and P M Njuho

14:45-15:00   13.   Production of genetically improved seed of subtropical and temperate
                    Eucalyptus and Pinus species for forest plantations in South Africa
                    W.R. Jones

15:00-15:15   14.   Early results from a Eucalyptus hybrid seedling versus clone trial
                    G.M. Galloway and T.K. Stanger

15:15-15:30   15.   Comparison between different yield stability procedures in maize (Zea
                    mays L.)
                    M.J.A. Alberts and C.S. van Deventer

15:30-16:00   Tea/Coffee

16:00-16:15   16.   Citrus rootstock breeding in South Africa: constraints, successes and
                    future prospects
                    Z. Bijzet

16:15-16:30   17.   Advances in controlled pollination techniques of Pinus species and
                    applications for commercial deployment
                    A. Nel, J. van Staden and A. Kanzler

16:30-16:45   18.   Application of chromosome doubling for the improvement of South
                    Africa’s indigenous flora
                    K. Hannweg and G. Visser

16:45-17:00   19.   Tailor-made deciduous fruit cultivars for South Africa
                    L.J. von Mollendorff
                                                                                        25


                             Session 3B Biotechnology
                           Chair: Prof Renée Prins

14:00-14:30   K4    GM Crops for Africa
                    Jennifer Thomson
                    Department of Molecular and Cell Biology, University of Cape Town

14:30-14:45   20.   Fibre Genomics: Assisting tree breeders to improve fibre quality in
                    forest plantations
                    Z. Myburg

14:45-15:00   21.   Horticultural Genomics: where will it take us?
                    D.J.G. Rees

15:00-15:15   22.   Application of proteomics as a tool to study plant protein expressions
                    and their responses to biotic and abiotic cues
                    B.K. Ndimba, S. Chivasa, W.J. Simon and A.R. Slabas

15:15-15:30   23.   Characterisation and identification of Sorghum Proteomes
                    R. Ngara, P.N. Ndabambi, D.J.G. Rees and B.K. Ndimba

15:30-16:00   Tea/Coffee

16:00-16:15   24.   Agricultural Biotechnology in South Africa
                    S.G. Mundree
                    CEO, PlantBio, The National Innovation            Centre   for    Plant
                    Biotechnology

16:15-16:30   25.   Current developments with regard to genetically modified organism
                    regulations
                    C.D. Viljoen

16:30-16:45   26.   The impact of pollen movement on Identity Preservation of maize
                    L. Chetty and C.D. Viljoen

16:45-17:00   27.   Detection of GMO in food products in South Africa: Implications of
                    GMO labeling
                    G.M. Botha, C.D. Viljoen and B.K. Dajee


17:30         Annual General Meeting
20:00         Dinner at Boesmanland
                                                                                           26


                             WEDNESDAY 15 MARCH 2006
                              Chair: Dr. Francois Koekemoer

07:30-08:00   Registration

8:00-8:30     K5    SAPBA – Invited speakers
                    Plant water-use efficiency and carbon isotope discrimination
                    Graham Farquhar
                    Environmental Biology Group, Research School of Biological
                    Sciences, Canberra, Australia

8:30-9:00     K6    The global need for a sustainable agricultural model
                    Roberto A. Peiretti
                    CAAPAS President (American Confederation of farmers
                    Organizations for a Sustainable Agriculture), Cruz Alta, Cordoba
                    Province, Argentina Republic


                                 Session 4 Biotic stresses
                                 Chair: Dr. Deidré Fourie

9:00-9:30     K7    Perspectives on breeding cereals for rust resistance
                    Zakkie Pretorius
                    Department of Plant Sciences, University of the Free State

9:30-9:45     28.   Will the new virulent race of Puccinia graminis tritici in Eastern
                    Africa threaten the South African wheat market?
                    A.J. Lesch and F.P. Koekemoer

9:45-10:00    29.   Marker-assisted breeding for durable stem rust resistance in wheat
                    R. Prins, Z.A. Pretorius, L. Herselman, C.M. Bender and M.J. Hayden

10:00-11:00   Tea/Coffee /Poster viewing

11:00-11:15   30.   Evaluation of the commercial utility of shortened forms of the Sr31
                    and Lr19 translocations
                    W.C. Botes, G.F. Marais, H.S. Roux and J.E. Snyman

11:15-11:30   31.   A host-pathogen study of stripe rust resistance in Triticum aestivum
                    L.J.Matthews, C.D. Viljoen, and R.L.V. Verhoeven

11:30-11:45   32.   Is Asian Soybean Rust a threat to common bean?
                    M.M. Liebenberg, A.J. Liebenberg and Z.A. Pretorius
                                                                                        27


11:45-12:00   33.   Genetic analysis of resistance to powdery mildew (Podosphaera
                    leuchotricha) in apple (Malus x domestica Borkh.)
                    Z. Simayi, M.M. van Dyk, S. Booi, R. Maharaj, M.C. Selala, M.K.
                    Soeker, I.F. Labuschagné and D.J.G. Rees

12:00-12:15   34.   Genetic analysis for resistance to Woolly Apple Aphid in apple
                    rootstock breeding populations
                    M.C. Selala, S. Booi, R. Maharaj, M.K. Soeker, M.M. van Dyk, Z.E.
                    Simayi, I.F. Labuschagné and D.J.G. Rees

12:15-12:30   35.   Genetic analysis of resistance to apple scab (Venturia inaequalis) in
                    apple, (Malus x domestica Borkh)
                    R. Maharaj, T Koopman, S. Booi, Z.E. Simayi, M.K. Soeker, M.C.
                    Selala, M.M. van Dyk, I.F. Labuschagné and D.J.G. Rees

12:30-12:45   36.   Studies on apple disease resistance genes
                    J. Mafofo and D.J.G. Rees

12:45-13:00   37.   Quantification of resistance to Meloidogyne incognita race 2 in local
                    soybean germplasm
                    H. Fourie, A.H. Mc Donald and D. De Waele

13:00-14:00 Lunch

                         Session 5A Breeding for quality
                           Chair: Prof Maryke Labuschagne

14:00-14:15   38.   Exploiting niche markets in the South African processing industry
                    F.P. Koekemoer, J.D. Theunissen and P. Graham

14:15-14:30   39.   Genetic mapping of fruit quality traits in apples (Malus x domestica
                    Borkh.)
                    M.K. Soeker, S. Booi, R. Maharaj, M.C. Selala, Z.E. Simayi, M.M.
                    van Dyk, I.F. Labuschagné and D.J.G. Rees

14:30-14:45   40.   The use of HPLC in predicting breadmaking quality
                    E. Koen, M. Labuschagne and T. Dessalegn

14:45-15:00   41.   Analysis of the proteome of red and green leaf phenotypes of ‘Bon
                    Rouge’ pear trees Pyrus communis. L. by 2-dimensional gel
                    electrophoresis.
                    M.J. Sehata, D.J.G. Rees and M.G. du Preez

15:00-15:15   42.   Four year results from a E. grandis composite Breeding Seed Orchard
                    G. J. van den Berg and T. K. Stanger
                                                                                     28



15:15-15:45   Tea/Coffee

15:45-16:00   43.   Genetic analysis of red pigmentation in Bon Rouge pears.
                    S. Booi, M.M. van Dyk, Z.E. Simayi, R. Maharaj, M.C. Selala, M.K.
                    Soeker, M.G. du Preez and D.J.G. Rees

16:00-16:15   44.   An investigation of the molecular mechanism underlying the
                    production of anthocyanin in 'Bon Rouge' (Pyrus communis, L) pear
                    trees and their green reverted sports.
                    M.G. du Preez, and D.J.G. Rees.

16:15-16:30   45.   Implementation of a robotics system for high throughput marker
                    assisted selection in the breeding of commercially important crops
                    such as apples and pears
                    L.H.H. Hüsselmann and D.J.G. Rees

                           Session 5B Genetic resources
                           Chair: Dr. Danie Theunissen

14:00-14:15   46.   Developing sustainable seed supply systems for Africa
                    R.B Jones and C. Dominguez (ICRISAT)

14:15-14:30   47.   Genetic diversity among traditional Ethiopian highland maize
                    accessions assessed by SSR markers and morphological traits
                    Y. Beyene, A-M. Oberholtster and A.A. Myburg

14:30-14:45   48.   Phylogeography of Eucalyptus urophylla based on the chloroplast JLA
                    region
                    K.G. Payn, F.M. Maleka, W.S. Dvorak and A.A. Myburg

14:45-15:00   49.   Fingerprinting SA cactus pear (Opuntia spp) germplasm using AFLPs
                    B.K. Mashope, L. Herselman, and M.T.Labuschagne

15:00-15:15   50.   Characterization of vernonia (Vernonia galamensis var. ethiopica) as
                    alternative industrial oil crop in Limpopo Province
                    S. Hussein, P. Mashela and A. Hugo

15:15-15:45   Tea/Coffee

15:45-16:00   51.   Apple cultivar and rootstock development for the South African
                    Second Economy: Enhancing economic and environmental
                    sustainability
                    I. F. Labuschagne
                                                                                    29


16:00-16:15   52.   Comparison of morphological and AFLP genetic diversity analysis of
                    Malawian cassava germplasm
                    I.R.M. Benesi, M.T. Labuschagne, L. Herselman and N.M. Mahungu

16:15-16:30   53.   Sweetpotato (Ipomea batatas (L) Lam) cultivar mixture by farmers in
                    Malawi: the basis for longer household food security
                    F.P. Chipungu, Aggrey J.D. Ambali and Nzola M. Mahungu

20:00         Greek evening at Marina terrace



                           THURSDAY 16 MARCH 2006

07:00         Breakfast
              Departure of delegates
                                                                                           30

                                       POSTERS

P1    Differential response of fourteen sunflower inbred lines to a rust (Puccinia helianthi)
      bulk isolate
      G.Chigeza, T. Debeila, and M.M. Liebenberg

P2    Effect of different environments on oil composition in high and mid-oleic sunflower
      varieties
      R. Coetzee , A. Hugo and M.T. Labuschagne

P3    Conventional breeding of high quality, seedless, Mandarins in the Eastern Cape
      N. Combrink, Z. Bijzet, and K Hannweg

P4    Comparison of AFLP and SSR markers for coffee (Coffea arabica L.) genetic
      diversity analysis
      Y. Dessalegn, L. Herselman and M.T. Labuschagne

P5    Association of bean caffeine content with cup quality and green bean physical
      characteristics in coffee (Coffea arabica L.)
      Y. Dessalegn, M. Labuschagne, G. Osthoff and L. Herselman

P6    Identification of root-knot nematode resistance in commercial maize material
      H. Fourie, A.H. Mc Donald & L. Ngobeni

P7    Using in vitro technology for citrus cultivar improvement
      K. Hannweg, Z. Bijzet, N. Combrink and G. Visser

P8    New sweet potato cultivars for resource-poor farmers
      S. M. Laurie, M.D Magoro, M.M. Mtileni, and A.A. Van den Berg

P9    Development of a new Ur-11 marker in dry beans
      L.A. Madubanya, M.M. Liebenberg and C.M.S. Mienie

P10   Alternative early generation selection methods to predict better malting quality in
      barley (Hordeum vulgare)
      A.F. Malan, H A Smit, R Oelofse and M Ncala

P11   Quality Protein Maize (QPM): facts and figures
      K. Mashingaidze

P12   Inhibitory activities of two apple polygalacturonase inhibiting proteins (MdPGIPs)
      L.B.T Matsaunyane, D. K. Berger and D. Oelofse

P13   Halo blight resistance in host differential cultivar ZAA 12 is conditioned by three
      genes
      P. Miklas and D. Fourie
                                                                                        31



P14   Host suitability of dry bean germplasm to the root-knot nematode species M. javanica
      H. Muedi and H. Fourie

P15   Purification of apple polygalacturonase inhibiting proteins 1 and 2 from transgenic
      tobacco, and an investigation into their inhibitory activities against fungal
      polygalacturonases
      D. Oelofse, I.A. Dubery, R. Meyer, I. Gazendam, G. De Lorenzo and D.K. Berger

P16   Studies on the S-locus and self-incompatibility in apple
      B. Okkers, J. Mafofo, I. Labuschagné and D.J.G. Rees

P17   DNA fingerprinting of table grape cultivars
      R. Prins, C.J. van Heerden, A.L. Burger, P. Burger and W.A. Smit

P18   Analysis of QTLs affecting dormancy release in apple (Malus x domestica Borkh.)
      M.M. van Dyk, M.C. Selala, Z. Simayi, S. Booi, R. Maharaj, M.K. Soeker, I.F.
      Labuschagné and D.J.G. Rees.
                                                                                             32

                                    ABSTRACTS


                                       Keynote 1.
                                  Monsanto Invited speaker


             Breeding for Performance under Drought; from Art to Science.

                                    Maarten van Ginkel

Department of Primary Industries, Victoria, Australia; Molecular Plant Breeding Cooperative
                                     Research Center

Breeding crops for performance under drought has received a new urgency with proof of
global warming now widely accepted in the scientific community. The record of breeding for
higher more stable yields under stress has overall been impressive, and not as stagnant as
some have claimed. Nevertheless, the future scenario will likely include not just expansion in
low rainfall areas, but also more erratic weather patterns. Demands on varieties will broaden.
Breeders are increasingly using some form of delineation of diverse production environments
into breeding domains, with certain traits only selected for certain domains while not for
others. This has allowed focus, flexibility and greater genetic gain. This situation will likely
change: increasingly over years past ‘fixed’ agro-ecological domains will begin to overlap,
with ‘freak’ weather extremes increasing in frequency. Varieties with good performance
under drought will need high levels of disease resistance and lodging tolerance for some
wetter years. Quality parameters will fluctuate with yield, while the market increasingly
demands uniformity, and is fully unappreciative of the challenge this places for breeders and
farmers. Farm-gate prices are not projected to increase for the major crops, at least in ‘free’
market economies. Hence the question is how to derive more stable production out of a less
predictable growing environment. In practise choice of crop, rotation, management and
marketing strategies become fluid and science seems to offer less a solution than in the past.
        What are approaches to address this conundrum of complexity? Three come to mind:
from data to information, from phenotype to genotype, from guessing to simulation to
prediction.
- On average less than 1% of the crosses that breeders make result in commercially relevant
varieties. Obviously there is room for improvement. The balance of breeding as an art and a
science, while a seemingly pleasing truism, needs to be shifted further towards ‘science’.
Many breeders have stacks of adaptation trial data in their computer Excel files or on their
shelves, analysed only for single locations or years. Modern GxE analysis has become very
powerful. It allows varieties best adapted over years to certain environments to be grouped
and identified. More recently the traits within varieties can be dissected and similarly
grouped, and likewise the weather and edaphic components of the locations. This provides
powerful insights into the specifics of ‘adaptation’, both in a temporal and spatial sense. It
allows the target population of production environments (TPE) to be appropriately weighed
during the breeding process. Breeders should work with statisticians turning the data obtained
from expensive trials into information.
                                                                                            33


- To the surprise of many some traits considered ‘quantitative’ are successfully being de-
constructed using Ockham's Razor. Genomic regions explaining components of such an
ephemeral trait as ‘yield’ are being located. Even performance under drought is being broken
down to its constituents (e.g. boron tolerance, nematode resistance), many driven for a large
part by a few genes with major effects. News is not all good: some traits have myriad
seemingly unpredictable expression states and still appear quicksilver. However, marker-
assisted selection is now a real tool that breeders at their jeopardy scoff at, side step or
neglect. Rather breeders should join biotechnologists in developing and studying the most
relevant marker populations. Exploitation of desired genes/alleles buried in wild crop
relatives associated with ‘linkage drag’ now seems more possible with marker approaches.
- Epistasis is the rule, rather than the exception. Not only do alleles between loci interact,
more so do their primary proteomic and secondary metabolomic products. This should come
as no surprise to any biologist. How to see the trees through the forest: play out genetic and
breeding alternatives using genetic simulation. Computers, while generally not ‘intelligent’ in
terms of self-learning, can compute complex relationships at high speed and low cost. The
relevant importance of certain genes and epistatic networks of multiple genes can be
modelled with simulation software and subjected to pilot selection processes without the need
for expense field trials at first, until the most promising options have been narrowed down.

maarten.vanginkel@dpi.vic.gov.au
                                                                                                34


                                           Keynote 2.

  Water quality of the lower Vaal River catchment and its influence on soils and crops

C.C. du Preez, M.G. Strydom, P.A.L. le Roux, J.P. Pretorius, L.D. van Rensburg and A.T.P.
                                        Bennie

Department of Soil, Crop and Climate Sciences, University of the Free State, P. O. Box 339,
                            Bloemfontein 9300, South Africa

The water of the lower Vaal River and its tributaries like the Harts, Modder and Riet Rivers is
mainly used for irrigation. An estimated area of 110 000 ha with soils ranging in texture
from sand to clay is either flood or sprinkler irrigated for cropping. Based on area the most
important crops in descending order, are wheat, lucerne, maize, groundnuts and soybeans.
Periodic reports have been received from irrigators of saline irrigation water causing crop and
possibly soil damage. These reports motivated this study.

The conductivity (EC) of irrigation water is a reliable indicator of salinity. The long-term
average EC of the Vaal River water increases from 52 m S-1 just below the Bloemhof Dam to
72 mS m-1 just above the confluence with the Orange River. This increase can be attributed
to accretions from the Harts and Riet Rivers which have poor quality water with EC values
above 115 mS m-1. The large fluctuation in water quality as a result of variations in
streamflow is a matter of concern. The EC of water in all these rivers increases from a
minimum at the end of the rain season (February to April) through the dry winter months to
reach a maximum at the beginning of the next rain season (August to October). In relatively
dry periods (1982 to 1984 and 1992 to 1994) the EC was generally high, with lower values in
relatively wet periods (1975 to 1976 and 1987 to 1988). Trend lines fitted through mean
annual EC values showed that the salinity in all rivers of the catchment will increase in
future. This will have without any doubt a negative impact on soils and crops.

Measurements in the catchment revealed that irrigation resulted in salinisation of some soils.
The salt accumulation was determined by soil type (texture, depth, internal drainage and
initial salt content), quality of irrigation water, type of irrigation system (flood or sprinkler),
and management practices (irrigation scheduling and leaching fraction). Relationships
between water salinity and crop yield as reported in literature were used to calculate the
influence of water quality on crop production in the catchment. Results indicated no yield
depression when the best water quality is used for irrigation. A reduction in the yields of
most crops was predicted when using the worst water quality for irrigation.

Information of this nature on water, soils and crops could be used with great benefit by plant
breeders to develop crops that are more tolerant to salts.

dPreezCC.SCI@mail.uovs.ac.za
                                                                                             35


                                          Keynote 3.

           Climate change and possible effects on agriculture in South Africa

                                     Johan van den Berg

               Business Development: ARS. Private Bag X15, Brandhof, 9324


Climate change is defined as the long-term change in climate over decades to centuries and
even longer periods of time. It is a natural phenomenon since the dawn of time with relatively
short cold periods of great severity, interspersed with long intervals of warm periods. Climate
is driven by energy and any change in energy levels will also have an effect on climate.
Energy levels from the sun as well as the composition of the atmosphere of the earth
determine to a large extent the climate of the earth.

It is a fact that global warming is taking place but the exact effect on climate is not defined.
The contribution of long-term climate change towards the total variation in climate in South
Africa is insignificant compared to the inter annual variation but there are some climate
change trends evident, depending on the length of the data series analyzed. A lack of accurate
historic climate records is responsible for uncertainty about the real extent of changes in
climate and projected effect on agriculture.

Some climate change trends that are detected are lower total amounts of cold units
accumulated due to higher minimum temperatures, a shift in the timing of cold unit
accumulation, changes in heat unit accumulation, timing and amount of rainfall.

Johan_Van_den_Berg.ARS@ars.co.za
                                                                                         36


                                        Keynote 4.

         The impact of GM crops in South Africa and their potential for Africa

                                   Jennifer A Thomson

    Department of Molecular and Cell Biology, University of Cape Town, South Africa

    It has been estimated that by the year 2025, sub-Saharan Africa will have a cereal
shortfall of nearly 90 billion tons. That will be the case if we continue to use current
agricultural practices. Therefore, if we wish to feed our people we will have to improve our
productivity. One of the ways to do this is to plant genetically modified (GM) crops to
improve yield. At a meeting of sub-Saharan agricultural experts held under the auspices of
UNIDO some years ago the following GM crops were highlighted as being important:
    • Crops resistant to African viruses such as Maize streak virus (MSV) and African
        Cassava mosaic virus (ACMV)
    • Crops resistant to parasitic weeds such as Striga
    • Insect resistant (Bt) maize of local varieties
    • Decreases in mycotoxins in maize
    • Drought tolerant crops.
    To date scientists in South Africa have developed maize resistant to MSV and have
applied to carry out field trials. An international consortium is working on developing
cassava resistant of ACMV. The company BASF has developed a bred line of maize that is
resistant to the herbicide imazapyr. Field trials have taken place in western Kenya where
farmers planted seeds coated with imazapyr and compared the yields with non-resistant lines.
The results were extremely impressive and a further set of field trials is underway.
    Bt maize has been planted by many small scale farmers in South Africa and again the
results have been impressive. There is also some indication that the maize is less prone to
post-harvest fungus infection and there may be an associated decrease in the levels of
mycotoxins in the stored grain.
    A research group at the University of Cape Town is working on developing drought
tolerant maize. They have targeted the indigenous resurrection plant, Xerophyta viscosa as
the source of their genes. The first transgenic plants were tolerant to dehydration, high
temperatures and salt. Other groups are targeting crops such as millet and sorghum.
    Finally, another UCT group is working on the development of oral vaccines in tobacco.
The viruses being targeted are HIV and the human papilloma virus that causes cervical cancer
in African women.

jat@science.uct.ac.za
                                                                                            37


                                         Keynote 5.

              Plant water-use efficiency and carbon isotope discrimination

                                     Graham Farquhar

Environmental Biology Group, Research School of Biological Sciences, Australian National
                                     University

    Theory suggests that the carbon isotopic composition of plants could give valuable
information about their transpiration efficiency – the amount of carbon dioxide taken up per
unit water lost by the leaves. Most carbon has mass 12, while about 1% is the stable (non-
radioactive) carbon isotope (13C). For some time it has been recognised that the 13C/12C ratio
varied in plants, and that the ratio is always less in plant organic matter than in carbon
dioxide in the air.
    We developed equations to explain how this ratio varied and showed that conservative
plants should discriminate less against 13C than plants that are using water quickly (Farquhar,
O’Leary & Berry, 1982). We predicted that the carbon isotope ratio could be used to screen
plants for water-use efficiency (carbon dioxide assimilation divided by water used) and
discussed this possibility for wheat. The theory was confirmed by examining carbon isotope
composition, water use and photosynthesis in different plant species. We demonstrated that a
mutant tomato plant lacking the hormone abscisic acid had unusually low water-use
efficiency and therefore large carbon isotope discrimination, and that phenotypic reversion of
both characteristics occurred when the missing hormone was fed to the plant. In all these
diverse experimental systems the changes in carbon isotope ratio observed matched the
theoretical predictions.
    To apply these predictions to the selection and breeding of a major agricultural crop, we
collaborated with a CSIRO plant breeder colleague to demonstrate that the genetic and
drought effects on water-use efficiency of wheat plants were revealed in their carbon isotope
composition in the manner predicted (Farquhar and Richards, 1984). The findings have been
confirmed in many other species. Jointly with Richards and his CSIRO group the underlying
plant and crop physiology of wheat water-use efficiency was explored and documented. A
collaborative program of selecting genotypes for improved water-use efficiency on the basis
of carbon isotope discrimination led to the release of two wheat varieties for dryland
Australia (Rebetzke et al., 2002), ‘Drysdale’ (released in October 2002) and ‘Rees’ (released
in September 2003). These new varieties out-yield standard varieties under dry conditions,
and the technique is now being applied in experimental breeding programs elsewhere,
including cowpeas for west Africa and barley for north Africa.
    Scientists at the international wheat and maize breeding centre, CIMMYT, are now
involved, and with colleagues from the International Rice Research Institute we have
demonstrated genetic variation of water-use efficiency of rain-fed rice.
    We recently identified a gene, Erecta, with a major control of transpiration efficiency in
the model plant species Arabidopsis (Masle, Gilmore & Farquhar, 2005). The group also
identified orthologues in other species. This significant finding opens the way for further
progress in identifying and adapting crop plants to dry environments.
                                                                                           38


Farquhar, GD, O'Leary, MH and Berry, JA (1982) On the relationship between carbon
   isotope discrimination and the intercellular carbon dioxide concentration in leaves. Aust.
   J. Plant Physiol. 9: 121-137.
Farquhar, GD and Richards, RA (1984) Isotopic composition of plant carbon correlates with
   water-use efficiency of wheat genotypes. Aust. J. Plant Physiol. 11: 539-552.
Rebetzke GJ, Condon AG, Richards RA and. Farquhar, GD (2002) Selection for reduced
   carbon isotope discrimination increases aerial biomass and grain yield of rainfed bread
   wheat. Crop Science 42:739-745.
Masle J, Gilmore S R and Farquhar GD (2005) The ERECTA gene regulates plant
   transpiration efficiency in Arabidopsis. Nature 436, 866-870

graham.farquhar@anu.edu.au
                                                                                             39


                                          Keynote 6.

                  The Global Need for a Sustainable Agricultural Model

                                     Roberto A Peiretti

Since the beginning of agriculture, ten thousand years ago, soils were mostly managed by
doing some kind of tillage. Even the strategy was useful to accompany an ever increasing
human demand for agricultural products; the “cost” in terms of soil erosion and degradation
was too high and should be consider as “no longer affordable” by humanity. Also, the
increase of the total agricultural production achieved during this period was obtained utilizing
a combination of the only two known basic mechanisms for this purpose: “the expansion of
the agricultural area by converting ecosystems into agro-ecosystems and/or the achievement
of a higher level of productivity on those already converted”. To be able to properly satisfy
the human future demand for agricultural products and considering that the area of land
suitable for agriculture is limited on earth, an entirely new agricultural system is needed. The
new system has to be able to allow the simultaneous achievement of a higher level of
productivity and sustainability.
During the last half of the past century, a new agricultural model based on No Till and on the
principles of the MOSHPA (Modern Sustainable High Productive Agriculture) was
developed and started to be adopted. The new model was based on a systemic view and in the
consideration of all the spectrum of human knowledge without discrimination (ex.: “from
agro-ecology to biotechnology”) but always science -and not ideologies- was the main
referential pillar.
The new model constitutes a valid and realistic way to evolve the agricultural process toward
the new goals: “higher productivity with profits and with a proper level of sustainability”.
Several American Countries that belong to CAAPAS (American Confederation of Farmers
Associations for a Sustainable Agriculture) had strongly participated in the development,
promotion, permanent improvement and adoption of this new and more evolved way to
understand and carry out the agricultural process. This new human agricultural approach will
play a central role at the time of attempting to solve what Dr. Norman Borlaug (Novel Peace
Laureate 1970) states as one of the bigger XXI century Human Challenge: “ To be able to
keep increasing the agricultural output but at the same time conserving the natural resources”.

sdrob@idi.com.ar
                                                                                           40


                                         Keynote 7.

                   Perspectives on breeding cereals for rust resistance

                                       Z.A. Pretorius

       Department of Plant Sciences, University of the Free State, Bloemfontein 9300

Rust epidemics have been synonymous with small grain cereal production since shortly after
these crops were introduced to South Africa in the 17 th century. Historically, wheat
breeding focused strongly on resistance to stem rust, caused by Puccinia graminis f. sp.
tritici, resulting in interspecific crosses as early as 1912. Due to ecological differences in
production regions in South Africa, a wide variety of wheat genotypes are required. This
implies genetic diversity and dedicated breeding programmes for winter, intermediate,
dryland spring and irrigated spring wheat types, respectively. Disease resistance is,
furthermore, seen as secondary to agronomic and quality traits and a perception exists that
fungicides can be applied when needed. Wheat breeders are not familiar with the identity of
the rust resistance genes in their germplasm, some environments may not be conducive to
rust epidemics, and key races may be absent during selection. Quite often new varieties
contain single gene resistance or some are susceptible to one or more of the rusts at the time
of their release. Almost a century after research on the cereal rusts was initiated, South
African wheat production remains vulnerable to stem rust epidemics, with at least four new
races detected during the past six years. This vulnerability is, however, not restricted to
South Africa. The current global threat of stem rust race Ug99 to wheat cultivars carrying the
Sr31 gene provides evidence that international breeding approaches have also been reliant on
monogenic resistance. In addition to stem rust, the damaging nature of wheat stripe rust has
been confirmed in several epidemics and pathogenic adaptations since its introduction in
1996. At present four races of P. striiformis f. sp. tritici have been confirmed in South
Africa. A national strategy for cereal rust control, with particular emphasis on pathogen and
host resources, breeding for resistance and centres of expertise, is urgently needed. Such a
strategy should take into account advances made in understanding the molecular basis of
plant-pathogen interactions and application thereof in crop improvement.

Pretorza.sci@mail.uovs.ac.za
                                                                                             41


 1. The influence of nitrogen on polymeric and monomeric proteins and quality in hard
                                     and soft wheat

                  M.T. Labuschagne, G. Meintjes and F.P.C. Groenewald

   Dept. of Plant Sciences, University of the Free State, P.O. Box 339, Bloemfontein 9300

The relative quantity of specific proteins, protein subunits, as well as amount and size-
distribution of polymeric proteins may vary due to environmental conditions. The aim of this
study was to determine the effect of different nitrogen treatments on polymeric and
monomeric proteins in wheat. Two soft white biscuit wheat cultivars, a cracker wheat and a
hard red bread wheat were planted under irrigation in a randomized complete block design
with three replications for two years running at six different nitrogen treatments. SE-HPLC
was used to determine the amount of monomeric and polymeric proteins, and various quality
characteristics were measured after harvesting. The large and small SDS-extractable
polymeric proteins were not influenced by different N levels. There was a strong cultivar
influence, where some cultivars had a larger reaction to N treatments than others. The later
application (at flag leaf stage) of nitrogen did not increase the protein fractions. The total
amount of N given seemed to have a bigger influence on the protein fractions than the timing
of the fertilizer application. The lowest N treatment consistently gave the lowest flour protein
content value. There was a strong correlation between flour protein content and large
monomeric proteins.

LabuscM.SCI@mail.uovs.ac.za
                                                                                            42


       2. Breeding new stone fruit rootstocks with specific stress resistance traits

                                        Hannél Ham

Agriculture Research Council (ARC) Infruitec-Nietvoorbij, Private Bag X5026, Stellenbosch,
                                          7599.

The stone fruit industry of South Africa has been challenged during the last decade by
changes in climatic, market demand and consumer preferences, both locally and on the export
market. For stone fruit cultivars to perform optimally, a combination of correct orchard
practises as well as rootstocks is a key factor in competitive and successful farming.
Therefore, the stone fruit rootstock breeding programme concentrates on the breeding of new
resistant rootstock for stress conditions associated with water logging, drought, high salinity
soil, lime induced iron chlorosis and nematodes (ring and root knot). Special emphasis is
also on assisting second economy producers who are often farming under less than optimal
site conditions. New rootstocks are bred conventionally and then tested under controlled
conditions to determine the level of stress resistance. The most promising resistant selections
are tested in rooting and hortological trails to determine their compatibility with commercial
cultivars. Currently it takes up to 40 years to breed a new stone fruit rootstock. Methods are
also investigated to shorten this time period.

hamh@arc.agric.za
                                                                                            43


   3. Investigating drought tolerant Eucalypt species and provenances for northern,
                                   coastal Zululand


                               R. A.W. Gardner and T. Swain

       Institute for Commercial Forestry Research, P O Box 100281, Scottsville 3209,
                              Pietermaritzburg, South Africa

The Zululand coastal region is one of the most important plantation forestry areas in South
Africa contributing 20 % of the country’s hardwood pulpwood. Annual rainfall is the main
environmental variable affecting tree growth across the region. Although a large proportion
(central areas) of the existing commercial forestry belt comprises well-watered, highly
productive sites, the peripheral areas to the north and west receive less rainfall and are
drought-prone, providing less productive growing conditions.
 Due to certain inadequacies in the Eucalyptus species being planted on these marginal sites
at the time, in 1992 the Institute for Commercial Forestry Research (ICFR) embarked upon a
search for better adapted (based on drought and disease resistance and pulping properties)
eucalypt species and provenances for coastal Zululand. Since this time, twenty species and
forty provenances of Eucalyptus were evaluated in two ICFR site-species interaction trial
series in the region. From these trials, only four eucalypts, namely Eucalyptus argophloia, E.
longirostrata, Corymbia citriodora and C. henryi have emerged as promising alternatives for
hot, dry areas in coastal Zululand. Eucalyptus longirostrata and E. argophloia are the only
species which appear insensitive to soil conditions. On the basis of saw timber production
(merchantable wood volume per hectare), C. citriodora and C. henryi have good potential for
moderately dry sites. On the basis of fibre production (tons screened pulp per hectare), C.
citriodora and C. henryi have good potential for moderately dry sites, whereas E.
longirostrata has good potential for both dry and moderately dry sites.
During 2001, comprehensive provenance/ progeny trials of the two most consistent
performers in the site-species trials, namely E. longirostrata and C. henryi, were established
on the coastal plain. Seedlots from six provenances of E. longirostrata and five provenances
of C. henryi were established in trials at two sites. At the dry site, Nyalazi, on the basis of
plot basal area at four years after planting, significant differences were recorded amongst E.
longirostrata provenances, but no significant differences were recorded amongst families for
either species. However, in the case of both species, the top families outperformed all
commercial control seedlots and clones.

rawg@icfr.unp.ac.za
                                                                                            44


           4. Adapting ARC fruit breeding goals to address changes in climate

                 J.P. Human, H. Ham, W.M. Pieterse and I.F. Labuschagnè

     ARC Infruitec-Nietvoorbij, Private Bag X 5013, Stellenbosch, 7599, South Africa

Due to climatic changes experienced in the Western Cape of South Africa, the fruit breeding
program of the ARC Infruitec-Nietvoorbij had to adapt breeding goals. Aside from the direct
effects of climate changes on fruit tree production, the interrelationships between fruit trees
and their pests and pathogens are also affected. Breeding for low chilling requirements in
peach, nectarine and apple was added as goals and specific studies were performed to
investigate genetic variation between and within families and between individual selections
for chilling requirements. Breeding stone fruit rootstocks for better adaptation to stressful
environmental conditions include resistance against soil salinity, lime induced iron chlorosis,
nematode and drought resistance. Apple rootstock breeding aims toward development of
climatically adapted rootstocks with resistance against woolly apple aphids, crown and root
rot and powdery mildew. Colour development in pear breeding receives much attention to
breed new pear cultivars less sensitive for increased temperatures.

humant@arc.agric.za
                                                                                             45


  5. Estimates of heterosis and association of genetic distance with heterosis in durum
                        wheat under different moisture regimes
                  1
                      Solomon, K.F, 2Labuschagne, M.T. AND C. D. 2Viljoen.
   1
       ARC-Small Grain Institute, Private Bag X29, Bethlehem 9700, 2Department of Plant
          Sciences, University of the Free State, P.O. Box 339, Bloemfontein, 9300

Knowledge on the amount of heterosis and heterotic pattern of parental materials would
greatly facilitate the development of drought tolerant cultivars. To this effect, use of
molecular markers is believed to help identify the heterotic pattern and predict heterosis or F1
performance. In durum wheat, information on the amount of heterosis and association of
heterosis with genetic distance under contrasting moisture conditions is limited. The
objectives of this study were to evaluate the level of heterosis for grain yield and yield
components, and the prediction potential of AFLP and agronomic traits based genetic
distances to mid parent (MP) heterosis and specific combining ability percentage (SCA%)
under well-watered and moisture stress conditions. Parental genotypes with different
responses to simulated soil moisture stress were crossed in six by six half diallel mating
design. Parental genotypes and their crosses were evaluated for their responses to moisture
stress conditions in glasshouse. Some hybrid combinations showed significant level of MP
heterosis for grain yield, yield components and harvest index (HI). The level of heterosis was
often different between stress and well watered conditions. Genetic distance (GD) matrices
were computed based on AFLP analyses. Morphological distance (MD) was calculated based
on standardised agronomic traits measurements. Cluster analysis of the parental lines based
on agronomic performance under stress conditions was similar to cluster analysis result based
on AFLP marker profile. Correlation between GD and MD was significant only at stress
moisture conditions. GD was correlated with none of SCA% distance matrices, except for
kernel weight under both moisture conditions. The correlation coefficients between MD and
SCA% matrices were positive and significant only for kernel weight and HI under well
watered and stressed conditions, respectively. GD was correlated to none of the MP-heterosis
distance matrices under any of the treatment conditions. MD was correlated significantly and
positively only to the MP-heterosis matrix for kernel weight under well-watered conditions.
The results suggest that evaluation of the association between genetic distance and heterosis
should be carried out under both stress and well-watered conditions. Absence of association
between AFLP based distance and heterosis for yield and most agronomic traits implied
heterozygosity per se is not a good predictor of heterosis under both well watered and
stressed conditions.


FekybeluS@arc.agric.za
                                                                                             46


   6. Some aspects of experimental design in a 10-year old Pinus patula progeny trial.


                                          A. Kanzler

 Sappi Forest Research, Shaw Research Centre, P. O. Box 473, Howick, 3290, South Africa.

Progeny testing plays a major role in forest tree breeding, above all to identify parents with
good general combining ability. The main objectives of these trials is the selection of parents
by means of the data from these tests, known as backward selection and the estimation of
variance components in order to determine genetic parameters. Experimental design has an
important impact on the efficiency and precision of progeny trials. In this study, a number of
different experimental designs are assessed in a 10-year old Pinus patula progeny trial.
The field test included thirty six open-pollinated families planted either in 18 replications of
single tree plots or in three replications of six tree row plots. In addition, a number of
different experimental designs, randomized complete blocks and lattice designs, were
compared.
The statistical and practical efficiency of each of these designs is assessed in the context of
long term crops such as conifer trees where other aspects such as, between tree competition,
generation interval and juvenile-mature competition are compared and discussed.

arnulf.kanzler@sappi.com
                                                                                            47


      7. Recurrent mass selection as a means to pyramid resistance genes in wheat

              GF Marais, WC Botes, HS Roux, L Snyman and MM Zaayman.

    Department of Genetics, University of Stellenbosch, Private Bag X1, Matieland 7602

A field-scale recurrent mass selection programme that is based on a male sterility gene (Ms3)
and hydroponic culture of cut tillers is being developed to facilitate breeding for complex
(pyramided) disease resistance. The procedure is backed by simultaneous development of
marker-aided selection for select resistance genes. Major advantages compared to existing
strategies are its potential utility for sustained gene pyramiding without sacrifice of genetic
gain for yield, adaptation and processing quality; maximization of opportunities for crossover
and gene recombination and low operational costs. Duration of the breeding cycle is limited
to four years by making use of single seed descent propagation in the off season, thus
reducing the time from making of crosses to varietal release to 7 or 8 years. Provision is
made for continuous upgrading of the base population with new resistance genes in a manner
that does not undo previous genetic gain. The procedure is continuously being improved and
adapted for greater efficiency.

gfm@sun.ac.za
                                                                                           48


8. Mechanisation and computerisation of yield testing in a maize breeding programme

                             R. W. Dunlop and J. D. Rossouw

         Monsanto South Africa (Pty) Ltd, P O Box 7424, Petit, 1512, South Africa

Line development breeders spend large budgets and amounts of time developing lines for
new hybrid combinations in a maize breeding program. However, without an effective yield
testing network, that thoroughly evaluates all the new hybrids, in all the necessary
environments, all this money and effort would be spent in vain. Breeding methodologies and
technologies have evolved over time and most breeding programs use modern biotechnology
to produce better hybrids. Yield testing, too, has progressed, from being labour intensive and
relatively inefficient to a highly mechanized, computerized and efficient part of the breeding
program. Looking at the four main operations within a testing network being planting, note
taking, harvesting and data analysis, they have all changed over time and this paper gives an
overview of the advancements in these four areas within Monsanto. Planting has changed
from being a highly labour intensive hand-planting operation to using tractor-drawn precision
planters, note taking has become computerized using handheld computers, no longer the
pencil and paper operation it used to be. Harvesting has evolved from hand harvesting to
using computerized twin plot harvesters and data analysis has become almost totally
automated. Some future advancements are also discussed.

rob.dunlop@monsanto.com
                                                                                              49


  9. Where eucalypt tree breeding and biotechnology meet: an unfortunate case study!

                                        Tammy Swain

     Institute for Commercial Forestry Research (ICFR), Box 100281, Scottsville, 3209

Eucalyptus nitens is the most important cold tolerant eucalypt species (CTE) currently grown
by the forestry industry in the summer rainfall regions of South Africa. The species is
recognised as the most cold and snow tolerant of the CTEs in South Africa, has moderate
frost tolerance and good pulping properties. The breeding programme for this species at the
ICFR has been ongoing since the 1980s and includes several provenance/progeny trial series
established over a range of sites, encompassing different site and climatic conditions.

One such trial series comprised two trials established on the Highveld during 1992 at
Arthur’s Seat and Goedehoop, with the aim of testing two collections from Barrington Tops
in northern New South Wales in Australia and once collection from Badja in central New
South Wales. Material from New Zealand and several South African landraces were also
included, as well as selected seedlots from previous trials, two E. grandis x nitens hybrid
controls and one E. macarthurii species control. It was later discovered that the trial at
Goedehoop was planted incorrectly, with several trays of treatments being planted in reverse
order. This error was discovered due to the fortuitous inclusion of the different species
controls, which are easily identifiable, but the extent of the error was unknown in those
blocks that did not include controls.

As the cost involved in the establishment, maintenance and measurement of forestry trials is
huge, methods were investigated whereby the information held in this trial could be retrieved.
In an attempt to determine which plots were correct, foliar samples were collected from
known and unknown plots, and RAPD’s were used to try and identify whether treatments
were correctly placed. As not all results from the RAPD screening were conclusive, this
exercise was repeated several times with different material, until the researcher felt that there
was enough information to re-draw the trial map with certainty.

Once the trial map and database had been re-ordered, the trial data were analysed and showed
that although there were no significant differences between the Australian and New Zealand
provenances, there were significant differences between families. The top families
outperformed the bred E. grandis x nitens hybrid controls. These results are similar to those
obtained from the sister trial at Arthur’s Seat which was planted correctly, and thus increases
the researcher’s confidence in the results of the RAPD screening technique.

tammy@icfr.unp.ac.za
                                                                                              50


10. Computer modelling: a tool for comparing selection strategies in a simulated poultry
                  population and how this can be used in forestry

                  Jon De Guisti 1,2, Annabel Fossey1 and Carolyn Hancock1
    1
      School of Biochemistry, Genetics, Microbiology and Plant Pathology, University of
                                KwaZulu Natal, South Africa
   2
     Institute for Commercial Forestry Research, P O Box 100281, Scottsville, 3209, South
                                           Africa

There are a number of different methods of artificial selection, including: individual
selection, between family selection, within family selection, family-index selection and index
selection. One method of testing the effectiveness and accuracy of the different selection
methods is the use of computer models. Computer models can be used to simulate selection
strategies and to predict what strategy will be the most appropriate for the improvement of a
particular trait. This investigation compared the effectiveness of the above mentioned five
selection strategies using five computer models for four different traits in a chicken
population, namely, egg weight, egg production, age at first egg and body weight.

Firstly, each of these traits was selected for independently using the first four selection
methods and secondly, the traits were selected for in pairs using index selection. For all traits
family-index selection produced the best response to selection. It is, however, often not
economically and practically efficient to utilize this selection strategy. Individual selection
proved useful when selecting for traits with a moderate to high heritability (egg weight and
body weight), due to the fact that individuals are selected based on their own phenotypic
values. In contrast, within family selection and between family selection proved to be more
effective for traits with a low heritability such as egg production. Individual selection and
family-index selection, however, resulted in a very rapid decline in the standard deviation for
all four traits, whereas, between family selection consistently resulted in the slowest drop in
the standard deviation thereby producing better responses to selection in later generations.
The impact of correlations between these traits was evident from the results of index
selection. For example, egg production is negatively correlated with egg weight making it
difficult to gain a correlated response in both these traits simultaneously.

A future improvement of the models may involve the inclusion of the effects of both
inbreeding and the environment. Another very important future aim will be to broaden the
scope of this work so that predictions and important breeding decisions can be made about
other species especially species of forestry populations. Being able to predict or accurately
estimate aspects about forestry populations is an extremely useful tool as the growth of a tree
population is a long process that takes many years. Having a way of modelling this growth
will allow decisions to be made at an earlier point in time and speed up research in forestry.

jonathan@icfr.unp.ac.za
                                                                                                 51


   11. Update on the Multiple Population Breeding Strategy implemented for Acacia
       mearnsii (black wattle) in South Africa - three-year growth measurements

               S.L. Beck 1, J de Guisti 1, R. W. Dunlop1 and M. D. V. Resende2
   1
    Institute for Commercial Forestry Research, P O Box 100281, Scottsville, 3209, South
                                           Africa
 2
   Embrapa Florestas, Ministry of Agriculture - National Center for Forestry Research Caixa
                     Postal 319, CEP 83411-000, Colombo, PR, Brazil

Recently the Acacia breeding programme at the Institute for Commercial Forestry Research
changed its emphasis from improving bark yield and quality, to improving timber yield and
quality while maintaining acceptable bark quality. In 2002 a Multiple Population Breeding
Strategy was implemented to cater for these changes. Six sub-populations were established
across different sites in KwaZulu-Natal, each was determined by origin of seed, i.e. from
historical and current selections as well as from controlled crosses. Each sub-population was
established as a progeny trial with a seedling seed orchard adjacent to it. The management of
the seed orchards will be determined according to the performance of the families within the
progeny trials. By establishing two different trial designs at each site will also allow one to
test the impact of each design for selection and variance estimation. Each sub-population
progeny test includes nine commercial and genetic checks as entries. To keep the sub-
populations unrelated for future crosses, the commercial and genetic checks are not included
in the single-tree plot BSOs.
This is the first time that a series of trials of this nature has been planted as part of a breeding
strategy for black wattle in South Africa. Data from these trials are undergoing rigorous
analysis using REML and BLUP, in order to set up a suite of analyses that can be routinely
performed on these data as the trials are measured annually. This will also provide an
appropriate decision-making tool, to use to select individuals for future generations.


This paper gives an update on some early growth measurements coming out of the trial which
are showing great potential for genetic gain in future. These early gains have been
extrapolated into final volume estimates to see if the predicted gains are real and can add any
value to the wattle industry in the future. The authors believe that this strategy and the
analytical techniques employed will identify the best individuals and families to be used in
future nucleus breeding populations. Ultimately the benefits of this strategy will be reaped by
the growers who choose to use the seed emanating from this breeding programme.



sascha@icfr.unp.ac.za
                                                                                              52


     12. Developing a non-destructive analytical technique for pulp yield in Black Wattle
                                      (Acacia mearnsii)

                               R. W. Dunlop1 and P M Njuho2
 1
   Formerly, Institute for Commercial Forestry Research, P O Box 100281, Scottsville, 3209,
                         South Africa (now employed by Monsanto SA)
2
  School of Statistics and Actuarial Science , University of KwaZulu-Natal, Private Bag X01,
                             Pietermaritzburg, 3209, South Africa

Acacia mearnsii (black wattle) is an important South African commercial forestry species,
providing a source of high quality raw material (fibre) for both the domestic and international
pulp and paper industries. Compared with many Pinus and Eucalyptus species, there has been
very little research into the wood and pulping properties of black wattle. The ability to assess
pulp yield in a non-destructive manner, using near infrared spectroscopy (NIR), is vital from
a tree improvement perspective. Destructive sampling and analysis results in a loss of the
genotype, while also being very expensive and time consuming. In order to assess some of
the important characteristics that make the species desirable from a fibre perspective, this
study investigates growth characteristics, wood density and pulp yield of trees grown on
three different sites and from three different age classes. In general, physical characteristics
such as utilisable height and diameter at breast height of the trees differed between sites and
increased with age. This trend was not reflected in the pulp yield or wood density results.
Pulp yield measurements ranged from 52.61 to 59.91% across all sites and age classes,
which, when compared to the pulp yield from many other forestry species, is relatively high.
The NIR calibration developed to predict pulp yield was acceptable with R2 = 78.9 % and a
standard error of calibration of 0.79. This calibration was used to measure the pulp yield
variation within the trees selected for this part of the study. Pulp yield decreased from pith to
bark and from the base of the tree to the top of the tree, with the highest values recorded in
the bottom 20% of the stem. The within tree variation data for pulp yield was analysed to
identify the best position for non-destructive sampling, and a model was then developed to
predict whole tree pulp yield based on this sample, which was taken at breast height. Near
infrared spectroscopy can be used to analyse small samples of sawdust, enabling prediction of
whole tree pulp yield.

rob.dunlop@monsanto.com
                                                                                            53


 13. Production of genetically improved seed of subtropical and temperate Eucalyptus
               and Pinus species for forest plantations in South Africa


                                         W.R. Jones

 Shaw Research Centre, Sappi Forests Research, PO Box 473, Howick, 3290, South Africa

Production of commercial quantities of genetically improved seed is a key step in translating
breeding and development into realised gains in field operations for the traits of interest.
Commercial seed utilized in the forestry industry has over many years been derived from
collections in natural stands, suitable provenances through to seed from elite orchards with
full pedigree control. Seed orchards are developed to perform specific functions and produce
specific products. These include Land race orchards, Breeding seedling seed orchards,
Seedling seed orchards and Clonal seed orchards. Seed orchards are based on a number of
selected trees from a range of families within a number of provenances. These groups of
diverse individuals often exhibit a range of flowering periodicity that influences out-crossing
rates and results in a combination of full-sib, half- sib and selfed progeny due to a lack of
control of the male pedigrees. If there is a high degree of variation for general combining
ability among males this could lead to inbreeding depression and will reduce expected gains.

The breeding systems within each genus will dictate the management strategy to produce
high value commercial quantities of genetically improved seed. Eucalyptus species are
mostly insect pollinated (entomophily) with preferential out-crossing, which is reinforced by
a gene-controlled incompatibility mechanism that impedes or prevents selfing. Pinus species
on the other hand are wind pollinated (anemophily) with sufficient degrees of asynchronous
flowering between male and female reproductive structures to favour outcrossing. Orchard
production is quantified by what is known as an efficiency factor (ef) for Eucalyptus species
(ef =x⁄(x+y)× 100 where x=clean seed; y=chaff) and for Pinus species (ef=x⁄z× 100 where
x=clean seed; z=number of cones). The efficiency factor is a measure of successful ovule
conversion into viable seed. The complex interaction between environmental factors such as
altitude, rainfall, and site together with species, breeding value and flowering periodicity
contribute to the ultimate recovery of viable seed with a particular genetic worth.

wayne.jones@sappi.com
                                                                                             54


          14. Early results from a Eucalyptus hybrid seedling versus clone trial

                             Galloway, G.M. and Stanger, T.K.

         Sappi Forests Research, Shaw Research Centre, PO Box 473, Howick, 3290

Eucalyptus grandis and Eucalyptus urophylla are important species for Sappi in the
subtropical coastal area of KwaZulu-Natal. These two species combine well in hybrid
combination, and clones of inter-specific crosses have outperformed pure species in
commercial compartments. In addition, hybrid clones of these species are tolerant of the
fungal diseases that are prevalent in this area, while E. grandis is susceptible. Disadvantages
associated with clonal production is the length of time it takes to test them before they can be
deployed commercially (minimum time of 11 years) and if a clone becomes susceptible to a
pathogen, large areas of a single clone can be affected.

With the advent of new techniques for controlled pollination (CP), and managing trees in pots
as potted orchards it is becoming more feasible to do mass CP of tested parents which have
good general hybridising ability. Advantages of mass CP are that seedlings from these
crosses would be able to be tested in half the time it takes to test a clone. Seedlings would
have a greater genetic buffering capacity and so not be as susceptible to disease as a clone
would be. A trial testing open pollinated and control pollinated E. grandis x E. urophylla
hybrid seed with commercial clonal controls was established in August 2003. The seed for
this trial was obtained from two sets of mini hedges, one in Howick and the other at
KwaMbonambi. Each set of mini hedges were planted with identical clones, at an espacement
of 2m x 2m with three E. grandis clones and eight E. urophylla clones.

There were five treatments of CP seed as well as four GU commercial clonal controls and
two seedling controls, one commercial E. grandis bulk seedlot from an orchard in Clan and
one E. urophylla bulk seedlot obtained from Indonesia. Tree diameter at breast height (DBH)
was measured at 24 months, and basal area was calculated, this takes survival into
consideration. These results are discussed in the presentation. The top ranked treatment in the
trial was a commercial GU clonal control. Four seedling treatments outperformed the next
two commercial GU clones. An evaluation of the defects was also made in each treatment in
the trial, as well as a subjective evaluation of obvious hybrids in each seedling block.

Geoff.Galloway@sappi.com
                                                                                                55


  15. Comparison between different yield stability procedures in maize (Zea mays L.)

                             M.J.A. Alberts1 and C.S. van Deventer2
                   1
                   Monsanto South Africa (Pty) Ltd, Petit 1512, South Africa
  2
      Department Plant Sciences (Plant Breeding), University of the Free State, P.O. Box 339,
                               Bloemfontein 9300, South Africa

Twenty three maize hybrids consisting of fourteen ultra short to medium maturity, 111 to 124
RM relative maturity (RM days), and nine late maturing hybrids with, 125 to 134 RM, were
evaluated for genotype x environment interaction (GEI) and yield stability across 42
environments from 2001 to 2003. The objectives were to estimate the components of variance
associated with the first and second order interactions and to determine their effect and
secondly to compare several statistical yield stability procedures. The procedures compared
were (1) coefficient of variability (CVi); (2) mean ( X ); (3) stability variance ( σ 2 i); (4)
ecovalence ( W i ); (5) regression coefficient ( bi ); (6) deviation from regression ( S 2 i ); (7)
                                                                                         d
cultivar superiority measure ( Pi ); (8) variance of ranks (S1) and (9) AMMI stability value
(ASV). Analysis of variance (ANOVA) showed the main effects due to years, locations and
the first order interactions (year x location) as highly significant. The main effect for
genotype, first order interaction (genotype x locations), (genotype x year) and second order
interaction (genotype x locations x year) were also highly significant. The highly significant
interactions indicate that genotypes need to be tested in several years and locations in order to
select stable genotypes.
Spearman’s rank correlation coefficient between the stability parameters indicated that
Shukla’s stability variance ( σ 2 i), Wricke’s ecovalence ( W i ), Eberhart & Russell’s deviation
from regression ( S 2 i ), the non-parametric stability measure of Nassar & Hühn, (S1) mean
                    d
absolute difference of ranks and AMMI stability value (ASV) had a highly significant
correspondence over the three years of study. The ASV and Nassar & Hühn’s (S1) were not
significantly correlated. No significant rank correlation between Lin & Binns’s superiority
measure ( Pi ) and Finlay & Wilkinson’s procedure ( bi ) with the other procedures were
found. The last two procedures are not recommended for use on their own as a measurement
of yield stability.

martin.alberts@monsanto.com
                                                                                           56


    16. Citrus rootstock breeding in South Africa: constraints, successes and future
                                       prospects

                                          Bijzet, Z

                            ARC, P/Bag X11208, Nelspruit, 1200

Attempts at rootstock breeding in South Africa date as far back as 1927 with the quest for an
alternative rootstock to the then widely used Citrus aurantium. Conventional breeding
techniques were used. Various constraints such as sterility, self and cross incompatibility,
nucellar embryony and a long juvenile period were encountered. As a consequence to this
little to no information regarding genetic control of economically important traits were
accumulated. However, various cross combinations have been made since 1993 that are
currently being measured for their ability to impart good quality and production to a scion.
Disease, pest and salt tolerance screening of the F1 progeny can also now commence as these
seedlings have passed through the juvenile period. Specific successes have been achieved
with regard to the manipulation of the juvenile phase and trustworthy screening methods for
the various rootstock traits. Advances in molecular genetic techniques and tissue culture-
based manipulation of plants have yielded new opportunities for developing citrus rootstocks
tolerant to environmental stress caused by diseases, pests, soil conditions and even tolerance
to adverse climate conditions. These advances, such as somatic hybridization, embryo rescue,
marker assisted breeding and many more, their application and impact on the South African
citrus rootstock programme are discussed.

zeldab@arc.agric.za
                                                                                           57


 17. Advances in controlled pollination techniques of Pinus species and applications for
                                commercial deployment

                            A. Nel1, J. Van Staden2 and A. Kanzler1
          1
           Sappi Forests Research, Shaw Research Centre, PO Box 473, Howick, 3290
2
    Research Centre for Plant Growth and Development, School of Biological and Conservation
       Sciences, University of KwaZulu-Natal, Pietermaritzburg Campus, Private Bag X01,
                                       Scottsville, 3209

The influence of four types of isolation material and pollen viability on cone survival and
seed set during controlled pollination of Pinus patula was investigated. Very low cone-
survival figures for controlled pollination of P. patula have been reported by the forestry
industry in South Africa. This places a major constraint on progress that can be made with
breeding in this species. Studies were conducted over two pollination seasons in 1997 and
1998 and the effect of treatments was monitored during the cone development period and was
extended to a cone-analysis study after harvest. The type of isolation material used during
controlled pollination has a major impact on cone survival and seed set, while the role of
pollen viability and self-pollination in successful controlled pollination was demonstrated. A
pollen study determined that differences in morphology and response to temperature
treatments were apparent among seven Pinus species used in an inter-specific hybridisation
programme. The seven species included were: P. caribaea, P. elliottii, P. greggii, P. oocarpa,
P. patula, P. radiata and P. tecunumanii. Different species responded differently to
temperature treatments of dry-stored and re-hydrated germinating pollen. The application of
these techniques for commercial deployment of improved material is outlined.

Andre.Nel@sappi.com
                                                                                             58


    18. Application of chormosome doubling for the improvement of South Africa’s
                                  indigenous flora

                              Karin Hannweg and Gerrit Visser

  ARC-Institute for Tropical and Subtropical Crops, Nelspruit, Mpumalanga, South Africa

South Africa has the highest recorded plant species density in the world. A large number of
these species have only marginal commercial and social potential due to size constraints. To
realise the full potential of this genetic heritage normally requires expensive long-term
breeding programmes. Chromosome doubling (induction of autopolyploids) is a technique
that can negate these size constraints within a relatively short research period. This technique
usually results in an increase in cell volume and consequently an increase in the size of plant
parts.

Many of the induced autopolyploids are expected to have at least one of the following
characteristics which would result in the improvement, or development of new economically
important plants: larger tuber, rhizome or root size; increased fruit size; enhanced flower size
and/or colour intensity, improved drought tolerance, increased bio-mass; improved
photosynthetic capacity; larger and/or thicker leaves; dwarfism; increased secondary
metabolite production e.g. medicinal compounds.


An overview of some of the results obtained will be presented.

Karin@arc.agric.za
                                                                                              59


                19. Tailor-made deciduous fruit cultivars for South Africa

                                   Leon J. von Mollendorff

                           ARC Infruitec-Nietvoorbij, Stellenbosch

The Breeding and Evaluation Division of ARC Infruitec-Nietvoorbij strives to develop tailor-
made deciduous fruit cultivars and rootstocks with superior fruit characteristics for all its
national and international customers. This Division places particular emphasis on research,
designed to allow the South African deciduous fruit industry to remain competitive on world
markets. Research activities in this Division include all aspects of conventional breeding,
development and commercialisation of apples, pears, plums, peaches, nectarines, apricots,
table grapes, raisin grapes and rootstocks. Breeding strategies in the Division focus mainly
on mass selection with regard to fruit quality, resistance against some of the major pests and
diseases in South Africa, storage ability and shelf life, and production improvement.

The impact of cultivar development by this Division on the South African Deciduous Fruit
Industry is clearly reflected in the number of cultivars released to the Industry during the past
15 years, as well as the foreign exchange earned through exporting fruit of these cultivars to
countries abroad. Ten pome and 67 stone fruit cultivars have been released to the South
African Deciduous Fruit Industry. Stone fruit cultivars bred by ARC Infruitec-Nietvoorbij
currently represent about 70% of the foreign exchange earnings from stone fruit. The
Japanese plum breeding programme, in particular, is recognised as one of the best breeding
stone fruit breeding programmes in the world. Thirteen plum cultivars have been released
during the past 15 years. These cultivars have unique characteristics that vary in colour
(yellow, red and black) and time of ripening (from early to late in the plum season) and have
a superior eating quality (e.g. high sugar content amongst other).

The development, release and marketing of new deciduous fruit cultivars world-wide are
becoming an extremely competitive enterprise. To safeguard the ARC’s products and return
on investment, commercialisation of ARC Infruitec-Nietvoorbij cultivars is becoming an
important goal. Currently, a large number of international license agreements have been
signed in most of the deciduous fruit producing countries of the world. Specific steps to
effectively control the volumes, fruit quality and selling prices for ARC cultivars produced in
foreign countries have been taken. The ARC also has to ensure that their cultivars do not
compete with the South African deciduous fruit markets abroad. Methods currently used to
facilitate the outcome of above-mentioned goals will be discussed.

vmollendorffl@arc.agric.za
                                                                                            60


     20. Fibre Genomics: Assisting tree breeders to improve fibre quality in forest
                                     plantations

                                   Alexander A. Myburg

 Forest Molecular Genetics Programme, Department of Genetics, Forestry and Agricultural
          Biotechnology Institute (FABI), University of Pretoria, Pretoria, 0002

Their long generation times, highly outbred nature and physical size pose important
challenges to the genetic improvement of forest trees. Nevertheless, forest tree breeders have
been remarkably successful in the early domestication and improvement of forest tree
species, most of which are still only a few generations removed from wild relatives. Forest
tree research recently entered the post-genomic era with the completed sequencing of the
poplar tree genome. At the same time, rapid progress is being made in the development of
genomic research tools in other forest plantation species such as pines and eucalypts. This has
massively expanded the resources available to forest tree researchers to develop
biotechnology applications that will accelerate tree improvement. Wood fibre development is
an important biotechnology target in forest trees and is the focus of a new industry supported
research programme at the University of Pretoria. The Wood and Fibre Molecular Genetics
Programme is a joint research venture of Sappi, Mondi and the University of Pretoria aimed
at the molecular improvement of fibre quality in fast-growing eucalypt and pine plantation
species in South Africa. Genomic approaches are being used to identify and isolate genes
involved in wood fibre development and to assess the genetic variation in these genes in tree
breeding programmes. Early successes include the isolation of the cellulose synthase gene
family of Eucalyptus and the development of molecular markers for routine fingerprinting
and marker-assisted breeding of forest trees.

zander.myburg@fabi.up.ac.za
                                                                                          61


                    21. Horticultural Genomics: where will it take us?

                                     D. Jasper G. Rees


       Department of Biotechnology, University of the Western Cape, Bellville, 7535


The development of genomics resources in plants started with the sequence of the
Arabidopsis genome in 2000. In 2002 the draft sequence for rice was released and within
2006 we will see the draft sequence for poplar, the first tree genome. As the technology for
genome sequencing and analysis becomes faster and cheaper, we can expect in the next
decade to have genome sequence data, and accompanying resources, for many more genomes
from commercially important crops, including those for the major horticultural species. The
genome sequences will completely change the way we analyse the genetics of a crop species,
and how we approach crop improvement. New technologies for high throughput genetic
analysis, mutation analysis, gene silencing, transcriptomics (gene expression analysis),
proteomics (protein expression analysis), and metabolomics (analysis of metabolite profiles
and fluxes), coupled with comparative gene models, will create novel ways of analysing and
understanding the biology and genetics of our major crop species. With planning already
advanced for genome sequencing projects for grape and at least one of the Rosaceae genomes
(apple, peach or strawberry), the new options will be outlined and the implications for South
African horticultural plant breeding and research will be discussed.

jasper@mail.biotech.uwc.ac.za
                                                                                                           62


      22. Application of proteomics as a tool to study plant protein expressions and their
                              responses to biotic and abiotic cues

                      B.K. Ndimba1, S. Chivasa2, W.J. Simon3, A.R.Slabas2,3
 1
     University of the Western Cape, Department of Biotechnology, Private bag X17, Bellville
                                7535, Cape Town, South Africa.
  2
      Creative Gene Technology, Integrative Cell Biology Laboratories, University of Durham,
                                              UK.
  3
      School of Biological and Biomedical Sciences, University of Durham, South Road, DH1
                                      3LE, Durham, UK
We have used a proteomics approach to study Arabidopsis thaliana cellular (Ndimba et al.,
2005) and extracellular (Ndimba et al., 2003; Chivasa et al., 2002) proteins under various
conditions. This approach facilitated the first discovery of extracellular ATP’s role as a plant
cell viability regulator (Chivasa et al., 2005). We have applied a relatively novel two-
dimensional difference gel electrophoresis to study effects of drought and salt stresses
Arabidopsis expressed proteins. Arabidopsis thaliana cell suspension cultures have been
used to investigate the effects of salinity and hyperosmotic stress on plant cellular proteins.
By carefully monitoring the pH of the apoplastic medium, we show that 200 mM NaCl and
400 mM sorbitol treatments induce extracellular medium acidification in Arabidopsis cell
cultures, a typical response of plant cells to salt and hyperosmotic stress. Using 35S labelled
methionine and cysteine, we demonstrated that NaCl causes a transient suppression of de
novo protein synthesis, from which the cells recover within 4 hours. Changes in the
abundance of cellular proteins 6 hours post NaCl and sorbitol treatments were analysed by
two-dimensional difference gel electrophoresis (2D DIGE). Of a total of 2949 protein spots
detected on the gels, 266 showed significant changes in abundance across 5 independent
experiments. Using matrix assisted laser desorption ionisation time of flight (MALDI-TOF)
mass spectrometry we identified 75 salt and sorbitol responsive spots. These fall into 10
functional categories that include H+ transporting ATPases, signal transduction related
proteins, transcription/translation related proteins, detoxifying enzymes, amino acid and
purine biosynthesis related proteins, proteolytic enzymes, heat shock proteins, carbohydrate
metabolism-associated proteins and proteins with no known biological functions. Overall,
this presentation demonstrates that proteomics can be used in virtually all aspects of plant
molecular biology and plant breeding to study protein expressions associated traits of interest.

References:
Chivasa, S., Ndimba B.K., Simon, W.J., Robertson D., Yu, X-L., Knox, J.P., Bolwell, P., Slabas, A.R.
Proteomic analysis of the Arabidopsis thaliana cell wall. Electrophoresis, 2002, 23, 1754-1765.
Ndimba, B.K., Chivasa S., Simon, W.J., Slabas, A.R. Proteomic analysis of changes in the extra-cellular matrix
of cell suspension cultures induced by fungal elicitors. Proteomics 2003, 3, 1047-1059.
Ndimba, B.K., Chivasa, S., Simon, W.J., Slabas, A.R. Identification of Arabidopsis salt and osmotic stress
responsive proteins using two-dimensional difference gel electrophoresis and mass spectrometry. Proteomics,
2005, 5, 4185-4196.
Chivasa, S., Ndimba, B.K., Simon, W.J., Lindsey, K., Slabas, A.R. Extracellular ATP defines a novel
endogenous external metabolite regulating plant cell viability. Plant Cell, 2005, 17, 3019-3034.

bndimba@mail.biotech.uwc.ac.za
                                                                                             63


              23. Characterisation and identification of Sorghum Proteomes

                  R. Ngara, P.N. Ndabambi, D.J.G. Rees and B.K. Ndimba

 Department of Biotechnology, University of the Western Cape, Private Bag X 17, Bellville,
                                          7535

Sorghum, a crop native to Africa, is ranked the fifth most important cereal crop in the world
and second after maize in Africa. In addition to being an important source of many food
products, sorghum is also an efficient source of animal feed, fibre and biomass. Sorghum is
drought tolerant, surviving under hotter and drier environmental conditions that are least
suitable for most other crops. For this reason, sorghum is not only an important water
efficient food crop in semi-arid regions, but also a good model plant for studying drought
tolerance mechanisms in cereals. In this study, we aim to use proteomic tools to compare the
proteomes of different sorghum varieties, these studies being the initial steps in achieving our
overall project goal of identifying and characterising drought tolerance related sorghum
proteins. This endeavour is also encouraged by the recent announcement that sorghum’s
genome will be sequenced in 2006.

The field of proteomics is increasingly becoming the technology of choice for studying
differential protein expression between crop varieties and crops subjected to environmental
stress conditions. In our study, we have extracted total protein from leaf, stem and root of
Sorghum bicolor seedlings. Proteins have been separated by 2-D gel electrophoresis (2-D
PAGE). After 2-D electrophoresis, the separated proteins are visualised on gels using either
Coomassie and/or silver staining. More than 1000 protein spots are observed from leaf
protein extracts with a small subset of these being variable between “red sorghum” and
“white sorghum” varieties. Experiments are in progress to explore proteome changes in
different varieties under conditions of salt and osmotic stress. Ultimately, protein
identification will be performed by mass spectrometry and genomic database searching, and
this should identify genes that could be used as biomarkers in plant breeding for the
development of new drought resistant varieties. Some candidates of interest can also be used
to transform and improve tolerance of other drought sensitive cereal crops.

rngara@mail.biotech.uwc.ac.za
                                                                                           64


                      24. Agricultural Biotechnology in South Africa

                                  Sagadevan G. Mundree

CEO, PlantBio, The National Innovation Centre for Plant Biotechnology, PostNet Suite 201,
                      P.Bag X6, Cascades, 3202, KwaZulu-Natal


The National System of Innovation in South Africa aims to use technology transfer and
technology diffusion to raise the level of technical competence in SMME’s and to allow them
to compete successfully in a global economy that is driven by knowledge. It also aims to
utilise this process to achieve the following national development imperatives:
              •   Accelerated economic development
              •   Sustainable development for poverty alleviation
              •   Rural development
              •   Small business development
              •   Human resource development
              •   Black economic empowerment
              •   Infrastructure development

One could achieve this by establishing programmes to support the development,
commercialisation and application of new biotechnologies, make better use of value chains,
and the existing science and technology infrastructure and achieve better integration of these
into local economic and provincial growth and development strategies as they shift towards a
knowledge-based economy. In plant biotechnology, there is a particular commitment to
developing centres of excellence, especially in fields where South Africa is a world leader or
has the genetic resources to do so.

PlantBio is a national initiative funded by the Department of Science and Technology (DST)
to promote plant biotechnology initiatives. Founded on the development of clear centres of
excellence in the field of plant biotechnology within South Africa, PlantBio focuses on the
commercialisation of results in this field. For example, forestry, agricultural crop yield
enhancement (especially of timber, sugar and sub-tropical crops), deciduous fruits, tropical
fruit crops and floriculture are key crop and biotechnology foci. A primary objective of
PlantBio is therefore to secure the capital and resources needed to bring technology to
market, i.e. commercialisation of research results.

mundree@plantbio.co.za
                                                                                            65


  25. Current developments with regard to genetically modified organism regulations

                                        C.D. Viljoen

           GMO Testing Facility, University of the Free State, P.O. Box 339, 9300

The advent of genetically modified organisms (GMOs) in crops has the potential to benefit
food production, through agronomical improvements as well as enhanced traits such as
improved nutrition. However, the introduction of commercial GMOs has also resulted in a
plethora of regulations that have a direct impact on crop production. During 2003, South
Africa ratified the Cartagena Protocol on Biosafety and the Genetically Modified Organisms
Act, 1997 (Act No. 15 of 1997) is currently being revised to incorporate the required
provisions. Under the Protocol, certain requirements are made with regard to the release, use,
and transit of LMOs (living GMOs). In addition to this, many countries have already
introduced mandatory GMO labelling. Although considered to be unfair trade barriers, such
regulations are motivated by the ‘consumers’ right to know’. In South Africa, as with many
other countries, there is very little consumer awareness of GMOs. While mandatory GMO
labelling may ‘frighten off’ consumers from using genetically engineered (GE) food it could
also be used as a tool to educate the benefits of this technology. However, mandatory
labelling is certainly not a consideration while the majority of consumers in South Africa are
oblivious to GMOs. Thus in order to accommodate those consumers that require GMO
labelling, voluntary labelling through identity preservation is currently under consideration
by the National Department of Health. To this end, a South African National Standard, Part 1
of 3: an ‘IP system for the production, storage, handling and transportation of non-genetically
modified unprocessed agricultural products’ was approved for release during 2005, with Part
2 and 3 to follow during 2006. It is ironic that while GMOs are already subject to heavy
regulatory scrutiny, non-GMO production will soon follow, placing an additional burden on
crop production.

viljoecd.sci@mail.uovs.ac.za
                                                                                         66


         26. The impact of pollen movement on Identity Preservation of maize

                                L. Chetty and C.D. Viljoen

          GMO Testing Facility, University of the Free State, P.O. Box 339, 9300

Maize is a staple in Africa and production is expected to increase with an increasing world
population. With the advent of modern breeding, it has become possible to alter the genetic
composition of a variety in terms of agronomic and/or quality traits using GM (Genetically
Modified) technology. Thus, the management of seed production including GM seed is
imperative. Identity Preservation (IP) can be used as a management tool during seed
production to maintain seed purity of a variety and thus minimize adventitious commingling.
While 99% seed purity may be considered acceptable for agronomic traits, it may not be
acceptable for specialized traits such as pharmaceutical seed production or for organic seed
production. Thus, an important factor in IP is to minimize pollen-mediated gene flow
(PMGF) and be able to predict the impact of practical isolation distances on seed purity in
seed production. Although there is no published data on PMGF for Southern Africa, studies
in maize have determined that PMGF can occur up to 650 m from a pollen source. The
environment is thought to have a considerable impact on PMGF. The aim of this study was
to determine the potential PMGF (PPMGF) and PMGF for maize in South Africa over
different environments. Bt yellow maize and conventional white maize was planted in a spilt
field design at Delmas and Lichtenburg during 2003/2004. Pollen was trapped during
flowering at distance intervals and genotyped to determine PPMGF. At harvest, PMGF was
determined by out-crossing using phenotypic as well as genotypic markers. These results
indicate that PPMGF can occur up to 400 m under favourable weather conditions.
Furthermore, the extent of PMGF was high between adjacent yellow and white maize rows as
expected with a decrease to 1% at 25 m and an average of 0.36% up to the furthest row of the
trial at 81.6 m thereafter. These results highlight the importance of establishing practical
guides, for producers in South Africa, to minimize commingling resulting from PMGF.

1999211681@student.uovs.ac.za
                                                                                            67


27. Detection of GMO in food products in South Africa: Implications of GMO labelling

                          G.M. Botha C.D. Viljoen and B.K. Dajee

GMO Testing Facility, University of the Free State, P.O. Box 339, Bloemfontein 9300, South
                                          Africa

Genetically modified (GM) crops currently account for 29% of crop production worldwide.
Since 1997, South Africa is the only African county to commercially grow GM crops, yet
very little consumer awareness exists. In contrast to this, there is strong consumer opposition
to GM foods in the European Union (EU) and Japan. Despite lack of regulations to provide
for food labelling that allows for consumer preference in South Africa, many products carry
negative or positive labels with regard to genetic modification. Although GMO (genetically
modified organisms) labelling does not have any bearing on the safety aspect of GMOs, it
gives consumers a choice between GM and non-GM, allowing them to balance concerns of
morality and perceived risk as well as informing them of GM. During 2005 the GMO
Testing Facility tested off-the-shelf maize and soy products to determine the uptake of GM
food into the human food chain as well as the validity of “non-GMO”, “GMO-free” or
“organic” labels, on local as well as imported products. Of the 58 products selected and
sampled randomly, 44 tested positive for the presence of GM. Furthermore, of the 20
products with a GM related label, 14 tested positive for GM. These results demonstrate the
extent of GM in the human food chain in South Africa and highlight the need for effective
regulations to protect consumers against misleading claims. Thus guidelines will have to be
established for producers and consumers alike for the use of GM related food labelling.

bothagm@yahoo.com
                                                                                          68


28. Will the new virulent race of Puccinia graminis tritici in Eastern Africa threaten the
                             South African wheat market?

                            A.J. Lesch and F.P. Koekemoer
                     Monsanto South Africa, Trade street, Napier 7270


Stem or black rust, caused by Puccinia graminis f. sp. tritici, has historically caused
severe losses to wheat (Triticum aestivum). Its control for over 30 years through the
use of genetic resistance in semidwarf cultivars is a remarkable success story.
However, this situation has led to decline in research and breeding efforts for
resistance in many countries (Singh et al., 2005). Resistance in many cultivars relied
on the gene Sr31 located in the 1B/1R wheat-rye translocation. During 1999
susceptibility of CIMMYT germplasm was noted in Uganda and an increase in stem
rust was also observed in Kenya. The race isolated from these areas proved to be
virulent on Sr31 and was named Ug99 according to the country and year of origin.
Ug99 has since spread to all parts of Kenya and Ethiopia and is virulent to many
popular cultivars. This race is feared to migrate further to North Africa, and through
the Arabian Peninsula and the Middle East, West Asia and, eventually, to South Asia
as has recently been witnessed for a stripe rust race virulent on Yr9 (Singh et al.,
2005). In South Africa, due to the negative quality characteristics (sticky dough)
linked to the Sr31 gene, breeders started using alternative sources of stem rust
resistance e.g. the adult plant resistance gene Sr2. In order to screen local germplasm
for resistance levels against Ug99, a set of lines and cultivars was screened at Njoro
experimental station in Kenya. Although most of the lines were extremely susceptible
some showed acceptable levels of resistance. These lines will be used extensively to
increase resistance levels in Monsanto’s breeding material. Monsanto also ventured
in a doubled haploid shuttle program with breeders from Kenya in order to screen new
lines for stem rust resistance.

Driecus.lesch@monsanto.com
                                                                                           69


         29. Marker-assisted breeding for durable stem rust resistance in wheat

        R. Prins1, 2, Z.A. Pretorius2, L. Herselman2, C.M. Bender2 and M.J. Hayden3
                    1
                   CenGen, 78 Fairbairn St., Worcester, 6850, South Africa
2
   Department of Plant Sciences, University of Free State, P.O. Box 339, Bloemfontein, 9700,
                                        South Africa
3
  Plant Genomics Center, University of Adelaide, Hartley Grove, Urrbrae, SA, 5064, Australia

The appearance and anticipated spread of a pathotype of Puccinia graminis f. sp. tritici with
virulence for the Sr31 gene for resistance have renewed interest in breeding for durable
resistance to stem rust of wheat. The Sr2 resistance gene has generally been accepted as a
source of durable resistance to stem rust, in particular when combined with other resistance
genes. Sr2 is not expressed in seedlings and is best observed by low rust severities in adult
plants grown in the field. Although Sr2 is genetically associated with melanin pigmentation
of wheat stems and glumes, as well as seedling chlorosis at higher temperatures, these
phenotypes are not consistently expressed. To determine the value of molecular markers
developed for Sr2, with the aim of application and thus more efficient stem rust resistance
breeding in South Africa, a collection of wheat cultivars, with or without the gene, was rated
for seedling chlorosis in a greenhouse. DNA was extracted from these plants and amplified
with a reported tightly linked microsatellite (SSR) marker (gwm533) and the two sequenced-
tagged microsatellite markers (STM) that were derived from it. Similar to the Australian
genotypes, it was shown that the use of the SSR marker gwm533 is precluded in SA
genotypes by the presence of a non-Sr2 associated allele of the same size. The STM markers
reliably amplified the correct alleles in most local and Australian control lines. However, in
several instances Sr2-associated alleles were amplified in presumably non-Sr2 carrying SA
cultivars. Further marker and phenotypic validation is underway to determine whether these
markers are indeed diagnostic in all South African genotypes.

cprins@lantic.net
                                                                                           70


    30. Evaluation of the commercial utility of shortened forms of the Sr31 and Lr19
                                    translocations

                   W.C. Botes, G.F. Marais, H.S. Roux and J.E. Snyman

  Department of Genetics, Stellenbosch University, Private Bag X1, Matieland 7602, South
                                         Africa

The Sr31/Lr26/Yr9/Pm8 gene complex and Lr19 are extremely useful sources of resistance
but have not been used commercially in this country because of linked deleterious quality
effects. Truncated forms of each were developed that are free of associated effects. A
shortened Sr31 translocation was produced by A Lukaszewski while we developed a
shortened Lr19 translocation. Both original translocations were shown to be associated with
a yield advantage. It has not been determined whether the yield enhancing effects were
retained in the shortened forms. We have crossed the two shortened translocations and used
the F1 to develop doubled haploids and single seed descent inbred lines. Lr19 and Sr31
specific markers were used to characterize the lines and to identify a subset of 40 lines that
include 10 representatives of each of the four possible factorial combinations. Since the Rht-
B1b (from W84-17-Lr19-149) and Rht-D1b (from Pavon–Sr31) genes segregated in the cross
it was necessary to also classify the lines with respect to Rht genes. Only semidwarfs were
used in the trial; double dwarfs and tall plants were excluded. The material was grown in
four replicated trials planted at representative Western Cape localities. The data was
analyzed to determine whether a positive correlation can be made between the translocations
and yield, and whether there is any indication of interactions of the two sources of foreign
chromatin. Preliminary results from (2004 and) 2005 trails indicate yield enhancing effects
do exist, but due to the influence of genotype by environment (GE) interaction on such data
these trials will be repeated in 2006 to also sample seasonal effects.

wcb@sun.ac.za
                                                                                            71


            31. A host-pathogen study of stripe rust resistance in Triticum aestivum

                     Matthews, L.J.1, Viljoen, C.D.2 and Verhoeven, R.L.V.2
 1
     Monsanto, Vermeulen st, Petit 1512, 2Department of Plant Sciences, University of the Free
                            State, PO Box 339, Bloemfontein 9300

Stripe rust, caused by Puccinia striiformis f. sp. tritici is a destructive wheat disease.
Management strategies are focused on producing resistant cultivars to achieve durable
resistance. It is therefore important to understand the underlying basis for the way in which
resistance functions. The aim of this study was to characterize the effect of the YrSp gene on
the host-pathogen interaction using histological and molecular techniques. In a quantitative
analysis of spore germination on susceptible and resistant plants, it was found that no
significant differences were noted during the first 8 days post inoculation, suggesting that
YrSp mediated resistance is post-haustorial. Furthermore, P. striiformis f. sp. tritici is
characterized by the absence of an appressorium and the presence of multiple pseudo-SSVI
(substomatal vesicle initial), during infection. The results of the confocal laser scanning
microscopy (CLSM) indicate that of the fluorochromes tested, the orange G probe and Uvitex
2B have the most potential to distinguish between fungal and plant tissue. The YrSp gene
was mapped to the short arm of chromosome 2B in an F2 Kariega x Avocet/YrSp population
(confirmed in an F3 population), using AFLP and microsatellite markers. In addition, the
presence of QTLs (quantitative trait loci) for major genes, conferring additive adult-plant
resistance (APR) in Kariega, was also confirmed.

lindy.matthews@monsanto.com
                                                                                               72


                     32. Is Asian Soybean Rust a threat to common bean?

                      MM Liebenberg1, AJ Liebenberg1 and ZA Pretorius2
       1
       ARC-Grain Crops Institute, Private Bag X 1251, Potchefstroom 2520, South Africa
  2
      Department of Plant Sciences, University of the Free State, P.O. Box 339, Bloemfontein
                                      9300, South Africa

Asian soybean rust, caused by Phakopsora pachyrhizi, is known to occur on many legumes,
including Phaseolus vulgaris. The disease was first reported on soybeans (Glycine max) in
South Africa in 2001. In 2004, it was found on the common bean (Phaseolus vulgaris). In
2005 a trial was planted under conditions of heavy disease pressure to obtain information on
the possible threat of Asian soybean rust to the common bean, and to identify possible
sources of resistance. Ninety-one Ph. vulgaris entries from different genetic backgrounds
were planted in single rows, each flanked by a row of susceptible soybeans. Preliminary
greenhouse inoculations were also undertaken. Eight weeks after planting the disease was
well established on the Ph. vulgaris leaves, and defoliation of mature leaves due to infection
by P. pachyrhizi took place approximately 12 weeks after planting, although infection levels
were lower than on the soybeans. Expanding leaves were not visibly affected. Genotypes
containing various Ur-genes conveying resistance to common bean rust were not resistant to
P. pachyrhizi. Common bean plants 3 to 5 m from infected soybean exhibited only low
levels of Asian soybean rust, and those less than 1 km away were not affected. For
greenhouse inoculations, a settling tower using dry spores, or direct transference of spores
from infected soybean leaves, using an artist’s brush, were the most effective methods.
Pustules appeared gradually and considerably later on Ph. vulgaris than on soybeans, and
infection levels were lower. A modified version of the trial will be planted during 2006.

LiebenbergM@arc.agric.za
                                                                                             73


  33. Genetic analysis of resistance to powdery mildew (Podosphaera leuchotricha) in
                            apple (Malus x domestica Borkh.)

       Z. Simayi1, M.M. van Dyk1, S. Booi1, R. Maharaj1, M.C. Selala1, M.K. Soeker1, I.F.
                              Labuschagné2 and D.J.G. Rees1
          1
              Department of Biotechnology, University of the Western Cape, Bellville, 7535
  2
      Breeding and Evaluation Division, Agricultural Research Council, Infruitec-Nietvoorbij,
                                       Stellenbosch, 7599

Apple powdery mildew, caused by Podosphaera leuchtricha, is one of the major diseases of
the cultivated apple in the Western Cape. The damage caused by this pathogen causes losses
in fruit quality and yield. Furthermore, the sector is affected as costs are incurred on the
management of this disease. The application of fungicides is one of the strategies mostly used
to control the spread of the pathogen. Since consumers demand high quality fruit with no
harmful chemical residues, alternative approaches that minimize the reliance on chemicals for
the control of this disease must be developed. Marker-assisted breeding for the development
of new varieties that exhibit durable resistance to the fungus is the best and safest way to
reduce damage caused by powdery mildew. In this study co-dominant microsatellite markers
are used to construct a framework genetic map for a population of 104 individuals resulting
from ‘African Carmine’ x ‘Simpson’, which has segregating resistance to P. leuchtricha. In
this work the generation of a preliminary genetic map will be described, using microsatellite
markers. From this map the location of genes contributing to resistance to Podosphaera
leuchtricha will be analysed, and markers linked to these genes will be of importance in
future selections for mildew resistance in the ARC Breeding program, which in turn will lead
to the development of mildew resistant cultivars in the future.

zsimayi@mail.biotech.uwc.ac.za
                                                                                             74


 34. Genetic analysis for resistance to Woolly Apple Aphid in apple rootstock breeding
                                        populations

      M.C. Selala1, S. Booi1, R. Maharaj1, M.K. Soeker1, M.M. van Dyk1, Z.E. Simayi1, I.F.
                                 Labuschagné2 and D.J.G. Rees1
          1
              Department of Biotechnology, University of the Western Cape, Bellville, 7535
  2
      Breeding and Evaluation Division, Agricultural Research Council, Infruitec-Nietvoorbij,
                                       Stellenbosch, 7599


The woolly apple aphid (Eriosoma lanigerum Hausmn.) is a major pest in apple production in
the Western Cape Province, South Africa. Breeding resistant rootstocks might provide a
necessary and environmentally friendly method of control. This study aims to map genes for
woolly apple aphid (WAA) resistance using microsatellite markers. A mapping population of
96 individuals from a ‘Northern Spy’ x ‘Cox Orange Pippin’ cross was generated. 52
seedlings were multiplied by in vitro propagation, planted in replicates of three blocks in a
greenhouse (20-25 0C) and evaluated for susceptibility and resistance to WAA. A linkage
map is being constructed using microsatellite markers to analyse the complete mapping
population. Fluorescently labelled microsatellite markers were used in linkage map
construction, and these were multiplexed by PCR for high throughput data generation. The
computer programs Genotyper and JoinMap were used in linkage map construction. The
current linkage map and analysis of quantitave trait loci (QTLs) for resistance to WAA will
be presented. Remove ref to future work if no data presented. The identification of QTLs for
WAA resistance will provide the tools required for the marker assisted selection of disease
resistant genotypes in future rootstock breeding, providing the basis for the development of
new disease resistant rootstocks that have reduced requirements for pesticide or fungicide
treatment.

mcselala@mail.biotech.uwc.ac.za
                                                                                             75


 35. Genetic analysis of resistance to apple scab (Venturia inaequalis) in apple, (Malus x
                                     domestica Borkh)

 R. Maharaj1, T Koopman2, S. Booi1, Z.E. Simayi1, M.K. Soeker1, M.C. Selala1, M.M. van
                     Dyk1, I.F. Labuschagné2 and D.J.G. Rees1
          1
          Department of Biotechnology, University of the Western Cape, Bellville, 7535
  2
      Breeding and Evaluation Division, Agricultural Research Council, Infruitec-Nietvoorbij,
                                       Stellenbosch, 7599

Apple scab, caused by the fungus Venturia inaequalis, is a major problem in the apple
industry, with lesions developing on both fruit and leaves. Control of scab is either through
extensive fungicide use, or by the use of scab resistant cultivars. Vf is the major gene
responsible for apple scab resistance and is found in cultivars like Prima and Priscilla.
However, from analysis of scab resistance in mapping populations containing the Vf gene, it
is clear that additional genes control the resistant phenotype. The incorporation of other scab
resistance genes in addition to Vf would provide greater and more durable scab resistance in
new cultivars. In this work, a mapping population from a ‘Lady William’s’ x ‘Prima’ cross,
consisting of 192 individuals, has been categorized into four classes of
resistance/susceptibility to apple scab. Microsatellite (simple sequence repeats) markers are
used in this study and a linkage map is being constructed for this population, which will
allow the (quantitative trait loci) QTLs for scab resistance to be identified. With the
availability of more QTLs combined with Vf, it will be possible to use markers linked to each
of the QTLs to select for more desirable cultivars resistant to apple scab in a high throughput
marker assisted selection program. This will lead to the selection of seedlings highly resistant
to apple scab for future evaluation work. In turn, this will lead to the production of new
cultivars with enhanced disease resistance, leading to reduced chemical inputs in production.

rmaharaj@mail.biotech.uwc.ac.za
                                                                                            76


                       36. Studies on apple disease resistance genes

                            Joseph Mafofo and D. Jasper G. Rees

       Department of Biotechnology, University of the Western Cape, Bellville, 7535

The NBS-type (nucleotide binding site) resistance gene analogs (RGA) represent a very large
gene family, which are involved in many examples of disease resistance in plants. Using PCR
primers designed against conserved sequence motifs in the NBS-RGA family, a mixture of
DNA fragments were amplified in ‘Golden Delicious’ and ‘Anna’ apple genomes, and these
were then cloned and sequenced. 136 sequences obtained this way matched RGAs from other
plant species. These were then analysed against Arabidopsis and cloned disease resistance
genes from rice resulting in classification into two subgroups with 74 being assigned to the
CNL class and 62 TNL. More RGA sequencing work was done using DNA from ‘Anna’. All
the clones with sequenced RGAs were further PCR amplified and products were spotted on
nitrocellulose paper. Hybridisation to isolated total RNA from apple plants infected with
Venturia inequalis was done to generate expression profiles for candidate disease resistance
genes from the sequenced RGAs. The analysis of variation in expression levels of the
different RGA genes will provide an important tool for the mapping of candidate genes that
control disease resistance. These will then be of value in the application of genetic selection
methods in the apple breeding program for the development of novel durable resistant
selections with reduced requirements for fungicide use in production.

jmafofo@mail.biotech.uwc.ac.za
                                                                                          77


     37. Quantification of resistance to Meloidogyne incognita race 2 in local soybean
                                         germplasm
                      1
                          H. Fourie, 1A.H. Mc Donald and 2D. De Waele
 1
   Agricultural Research Council-Grain Crops Institute, Private Bag X1251, Potchefstroom,
                                    2520, South Africa
2
  Catholic University of Leuven, Laboratory of Tropical and Crop Improvement, Kasteelpark
                            Arenberg 13, 3001 Leuven, Belgium

Host plant resistance to Meloidogyne incognita race 2 is a useful and cost-effective tool for
optimisation of soybean yield as well as profitability of the crop. Locally no nematicide is
currently registered on soybean and most crops used in soybean rotations are also susceptible
to M. incognita race 2. Subsequently local and foreign soybean genotypes were evaluated for
resistance to this nematode species in greenhouse experiments. Local cultivar LS5995
exhibited resistance to M. incognita race 2 and had the lowest RF-values, number of eggs per
root system and number of eggs per g of roots of all genotypes evaluated. LS5995 also
consistently expressed root-knot nematode resistance under microplot and field conditions
since it maintained low populations densities of both M. incognita races 2 and 4, as well as
M. javanica. Furthermore, data on the quantitative relationships between soybean crop
performance and initial population densities of M. incognita race 2 obtained in hailnet cage
and microplot studies showed that LS5995 has a damage threshold level that is 10 times
higher than that of a susceptible standard. According to pre- and post-infectional and
histological studies resistance in cultivar LS5995 manifested in the inhibited development
and reproduction of M. incognita race 2. It was identified as post-infectional antibiosis
resistance. Two molecular markers were linked to M. incognita race 2 resistance in cultivar
LS5995. These markers are currently used in a marker-assisted breeding programme (MAS)
to expedite the breeding process.

FourieHD@arc.agric.za
                                                                                              78


          38. Exploiting niche markets in the South African processing industry

                      F.P. Koekemoer, J.D. Theunissen and P. Graham

                          Monsanto, PO Box 556, Bethlehem, 9700

The transition of the world’s wheat industry in the past ten years from a government driven,
strategic commodity to a consumer driven, profit motivated industry is redefining the
meaning of quality. As much as we might prefer the comfort of the known world of the past,
change is upon us. We are not faced with the question of whether to change, but whether we
are going to attempt to control the change, rather than be controlled by it. We can be part of
the problem, or part of the solution (Dryan, 2001). The South African wheat market cannot
be isolated anymore from what is happening in the rest of the world, therefore classification
of local wheat cultivars is an attempt to provide the South African wheat industry with new
cultivars that perform well agronomical and possess suitable and unique milling, rheological
and baking characteristics. Before the South African wheat market was deregulated in 1998
the wheat farmer received 40% of the total profit available and the processing industry 17%
after deregulation the situation was reversed and the farmer receives 15% while the
processing industry receives 42%. With the steadily annual rise of input costs and with the
wheat price which is also below import parity the economical production of wheat is getting
tougher every year. Wheat is a self fertilized crop and this makes farm saves seed (FSS) a
viable option to reduce input costs. Increased FSS is putting the profitability of wheat
breeding programmes under pressure. In order to change the scenario Monsanto developed a
niche market cultivar to get back some of the profits being made by the processing industry to
the advantage of the farmer and their breeding programme. This season 50 000 tons had been
produced from this line and because the quality characteristics of this line is so unique, it can
not be mixed with the main bread wheat stream. Therefore, a closed loop system had been
developed to protect the identity of this line. Niche market lines should not have a yield drag
otherwise the profitability of such a system is under pressure. The system developed by
Monsanto had been a long and hard learning curve which paves the way for other niche
products which are being in the process of development.

Francois.petrus.koekemoer@monsanto.com
                                                                                                    79


       39. Genetic mapping of fruit quality traits in apples (Malus x domestica Borkh.)

      M.K. Soeker1, S. Booi1, R. Maharaj1, M.C. Selala1, Z.E. Simayi1, M.M. van Dyk1, I.F.
                                Labuschagné2 and D.J.G. Rees1
          1
              Department of Biotechnology, University of the Western Cape, Bellville, 7535
  2
      Breeding and Evaluation Division, Agricultural Research Council, Infruitec-Nietvoorbij,
                                       Stellenbosch, 7599


Apple fruit quality is of utmost importance to apple farmers and breeders. This is reflected in
major international markets, where failure to meet specifications can result in shipment
rejections, reduced returns to growers and a damaged reputation as a supplier of top quality
fruit. Fruit size, colour, texture, firmness and taste are all traits that affect the quality of fruit.
In this study the genetic contribution of these traits is being evaluated in order to generate the
genetic markers required for the application of marker assisted selection in fruit quality
breeding. Two mapping populations ‘Prima’ x ‘Anna’ and ‘Golden Delicious’ (GD) x
‘Priscilla’, consisting of 95 seedlings and 120 seedlings, respectively, were used in the study.
Fruit samples were analysed using visual and sensory techniques. This data was correlated
using statistical programs. DNA was extracted from the progeny, and microsatellites were
amplified by PCR using published and predicted primers. Scoring of these marker alleles was
performed using Genotyper, and then JoinMap was used to construct the genetic map.
The location of quantitative trail loci (QTL) will be analysed using MapQTL. Comparative
genome analysis and the role of various genes on the outcome of fruit quality can then be
investigated, using the integrated genetic map, and the QTLs identified can be used for
marker assisted selection, to increase the speed and efficiency of the apple breeding program.

ksoeker@mail.biotech.uwc.ac.za
                                                                                             80


                   40. The use of HPLC in predicting breadmaking quality

                  Elizma Koen1, Maryke Labuschagne1, and Tadesse Dessalegn2
  1
      Department of Plant Sciences, University of the Free State, P.O. Box 339, Bloemfontein,
                                       9300, South Africa
              2
                Adet Agricultural Research Centre, P.O. Box 8, Bahir Dar, Ethiopia

The functionality of wheat flour is primarily determined by its proteins. Hence, considerable
effort has been made to fractionate and quantify these protein constituents responsible for
quality differences. HPLC is a powerful tool to study native protein aggregates and
physicochemical basis of baking strength, and allows the rapid assessment of baking quality
of wheat genotypes in breeding programmes. RP-HPLC separates proteins based on the
difference in surface hydrophobicity, which compliments techniques such as SE-HPLC,
which resolves proteins based on molecular size. Both methodologies accurately separate the
three main classes of endosperm proteins: glutenins (polymeric proteins), gliadins, and
albumins and globulins (monomeric proteins). This is particularly important because the
relationship between protein classes (e.g., glutenin-to-gliadin ratio) and the molecular size
distribution of polymeric proteins affect quality attributes. In this study 13 Ethiopian and two
South African bread wheat cultivars were compared to 15 Ethiopian durum wheat cultivars in
two diverse environments. This was done to assess the effect of HPLC determined storage
proteins on the breadmaking quality. It was found that across environments, protein fractions
had a major influence on quality. The amount of polymeric proteins in bread wheat was
significantly higher in the high protein environment, but in durum wheat the opposite was
found. Durum wheat had higher percentages of large polymeric and small monomeric protein
fractions, compared to tested bread wheat. In bread wheat the large monomeric proteins,
mainly gliadins, had a consistently significantly negative effect on quality. An increase in the
polymeric-to-monomeric protein ratio, lead to an improvement of quality characteristics.
These results show that RP- and SE- HPLC are well suited for analysis of cereal proteins. In
breeding and genetic studies HPLC has the potential to allow varietal identification, purity
and selection for specific characteristics.

koene.sci@mail.uovs.ac.za
                                                                                            81


41. Analysis of the proteome of red and green leaf phenotypes of ‘Bon Rouge’ pear trees
                 Pyrus communis. L. by 2-dimensional gel electrophoresis

               M. James Sehata, D. Jasper G. Rees and Marlene G. du Preez


       Department of Biotechnology, University of the Western Cape, Bellville, 7535

The production and stability of fruit skin colour of red and blush pears determines their
economic advantage over green or yellow-green skinned fruit. Red and blush pears often
lose their colour due to various environmental and post-harvest storage conditions. To
investigate the molecular basis of the stability and loss of fruit skin colour, we use a
proteomics approach to characterise the extreme differences found between 'Bon Rouge' and
'Bon Rouge' reverted pears. 'Bon Rouge' is characterised by red skinned fruit resulting from
anthocyanin production and was derived from a bud mutation of ‘William’s Bon Chretien’
(Bartlett). The production of red and green skinned fruit on the same tree presents a system in
which to study the control of colour development under the same set of environmental
conditions in an identical genetic background. We analysed a series of equivalent samples
from 'Bon Rouge' and reverted leaves and compared the proteome of the two phenotypic
states (red and green leaves) during colour development using 2-dimensional gel
electrophoresis. Our preliminary proteomic analysis shows substantial changes in the patterns
of protein expression that were further characterised by mass spectrometry. Our previous
analysis of changes in gene expression in this system implicates a range of stress response
proteins, and we aim to evaluate the relationship between these proteins and those observed
by 2-dimensional electrophoresis methods. Additionally, we will use quantitative RT-PCR to
measure the differences in expression of the genes identified by this proteomics approach.

jsehata@mail.biotech.uwc.ac.za
                                                                                           82


       42. Four year results from a E. grandis composite Breeding Seed Orchard

                           G. J. van den Berg and T. K. Stanger

 Shaw Research Centre, Sappi Forests Research, P.O. Box 473, Howick, 3290, South Africa

In tree improvement one type of multiple population breeding makes use of composite
Breeding Seed Orchards (cBSOs). Theoretically all the sub-populations of a species breeding
population are managed together in one series of cBSOs. Thus there will always be less
cBSOs required than Breeding Seed Orchards (BSO). Initially the best elements in all BSOs
would be included in each cBSO. As the selection cycle progress a nucleus of elite families
will start to emerge for the environment in which the cBSO is planted. Each time a cBSO is
regenerated, the best families from the other cBSOs in the series will be included, to
challenge the members of the nucleus.

In the early 1990’s a series of E. grandis BSOs were established in order to manage
genotype-environment interaction using the multiple population approach, and allowing seed
production for specific regions. Results from these trials showed that the genotype-
environment interaction for the BSOs in the KwaZulu – Natal midlands area were of no
practical importance. A decision was made to use the common core of the best families from
these E. grandis BSOs to establish an E. grandis cBSO. In December 2000 one of a series of
E. grandis cBSOs was established at the Clan plantation, KwaZulu-Natal. The main
objectives of this trial are to provide the KwaZulu-Natal midlands area with an elite seed
source, as well as to test the progeny from the selections made in the original BSO series. In
series with this cBSO, a progeny trial at Ixopo and a cBSO at KwaMbonambi were
established to confirm the lack of genotype-environment interaction and to conserve this
valuable genetic material.

Mid-rotation results form these trials confirm that no significant genotype-environment
interaction occurred for these families between the different sites. Significant differences
between families however, were detected. More than fifty percent of the families significantly
outperformed the control treatment; potential genetic gains can therefore be quantified.

Gert.vandenBerg@sappi.com
                                                                                           83


              43. Genetic analysis of red pigmentation in Bon Rouge pears

S. Booi, M.M. van Dyk, Z.E. Simayi, R. Maharaj, M.C. Selala, M.K. Soeker, M.G. du Preez
                                    and D.J.G. Rees

       Department of Biotechnology, University of the Western Cape, Bellville, 7535

Pome fruit cultivation and production is of major economic importance in the Western Cape
region of South Africa. 'Bon Rouge' is a red pear cultivar that was derived from a rare,
spontaneous bud mutation of the green pear cultivar William's Bon Chretien (Bartlett). In this
study we aim to map the genetic location of this event, with a long-term view to cloning and
characterising the gene involved. A closed cross between 'Bon Rouge' and 'Packham's
Triumph' pears generated an F1 population of 800 seedlings with a 1:1 segregation of the
red:green phenotype, indicating a simple Mendelian inheritance of this trait. We are currently
mapping apple and pear microsatellites on the F1 progeny. Both apple and pear microsatellite
markers were used in this study because of the high level of between-species cross-reaction.
Microsatellites that show a high level of polymorphism are being used to construct a linkage
map with the aim of identifying those markers that are tightly linked to the gene controlling
the red/green trait. This will provide the basis for future genetic mapping in other pear
mapping populations. To saturate our map in the region of the “red” locus, Amplified
Fragment Length Polymorphisms (AFLPs) will be used to obtain a high density map in the
region of this locus, which will provide the information required for a chromosome-walking
strategy. The project will increase our understanding of the molecular mechanisms of colour
development in pears, which will be of importance in the future for both breeding and
production of pears with the desired colouration.

sbooi@mail.biotech.uwc.ac.za
                                                                                             84


     44. An investigation of the molecular mechanism underlying the production of
  anthocyanin in 'Bon Rouge' (Pyrus communis, L) pear trees and their green reverted
                                         sports

                        Marlene G. du Preez, and D. Jasper G. Rees


       Department of Biotechnology, University of the Western Cape, Bellville, 7535

The production and stability of fruit skin colour in red and blush pears are desirable traits in
many export cultivars under commercial production in the Western Cape region of South
Africa. To investigate the underlying molecular mechanism controlling the production,
stability and loss of anthocyanin, we aim to characterise the extreme differences found
between red leaved 'Bon Rouge' (Pyrus communis, L.) pear trees and their green sports, using
a number of molecular tools. 'Bon Rouge' was derived from a rare, spontaneous bud mutation
of the green pear cultivar 'William's Bon Chretien' (Bartlett) and is characterised by red
skinned fruit resulting from anthocyanin production. 'Bon Rouge' was observed to revert at a
high frequency, producing green tissues in clonal stripes on stems and fruit. The production
of red and green skinned fruit on the same tree presents a system in which to study the control
of colour development under the same set of environmental conditions or variables in an
identical genetic background. Differences in gene expression between the two phenotypic
variants were measured by differential display and confirmed by quantitative RT-PCR. Seven
of the differentially induced cDNA clones showed significant similarity to genes in database
and include those associated with light stress, pathogenesis responses, and protein synthesis.
We compared the proteome of the two phenotypic states (red and green leaves) during colour
development using 2-dimensional gel electrophoresis. Our preliminary proteomics analysis
shows substantial changes in the patterns of protein expression that was further characterised
by mass spectrometry. HPLC profiles were generated for red and green plant tissues and
individual peaks characterised by 1- and 2-dimensional 1H and 14C NMR. One compound
was fully characterised as a natural anti-oxidant previously identified in tart cherries. Our
preliminary analysis suggests that many stress response genes are involved in anthocyanin
production in 'Bon Rouge' and we aim to assess whether the underlying mechanisms of
colour development in this cultivar can be understood and in the longer term, manipulated.

mgdupreez@uwc.ac.za
                                                                                             85


 45. Implementation of a robotics system for high throughput marker assisted selection
       in the breeding of commercially important crops such as apples and pears

                       Lizex H.H. Hüsselmann and D. Jasper G. Rees

        Department of Biotechnology, University of the Western Cape, Bellville 7535

The implementation of a high throughput system for genetic mapping and marker assisted
selection requires the use of automated technologies to increase throughout, improve
reproducibility and reduce the “burn-out” of staff. While PCR-based methods such SSRs
(microsatellites) are routinely performed in many laboratories, the screening of thousands of
samples for the desired gene is greatly enhanced by the use a high throughput robotics system
that can perform these routine tasks. The epMotion 5075 VAC is a robotic system that can be
used in various configurations to fully automate the liquid handling requirement of the MAS
program. The system is designed to make time-consuming tasks simple, reliable and
reproducible. Twelve positions are available in the work area, which accept every type of
microtest and PCR plate; racks for individual tubes and pipette tip boxes. The software
includes methods such as nucleic acid purification and allows for programming of bespoke
protocols, which are executed fully automatically in a short time. Pooling of four multiplexes
(96 samples each) into one plate is completed in less than one hour. Setting out of a PCR
plate with DNA samples (96) and PCR mastermix is done within 15 minutes. The
implementation of the epMotion 5075 as part of a semi-automated system for marker assisted
selection in the apple breeding program will be presented. It is expected that this will provide
the technology required to increase the throughput of the breeding program to 100 000
seedlings per annum, with a 95% elimination rate based on the genetic marker selection
applied.

lhusselmann@mail.biotech.uwc.ac.za
                                                                                             86


                46. Developing sustainable seed supply systems for Africa

                          Richard B Jones and Carlos Dominguez

      The International Crops Research Institute for the Semi-Arid Tropics (ICRISAT)

Improved seed of well-adapted crop varieties can benefit small-scale farmers by increasing
the value and productivity of assets at hand – be they land, labor or capital. The resulting
productivity and quality gains improve food and nutritional security, and increase rural
incomes. These combined benefits should drive the development of viable seed systems and
yet most small-scale farmers in sub-Saharan Africa continue to rely on indigenous seed
systems.
The liberalization of seed markets during the past decade or so has encouraged international
seed companies to increase their stake in the market. A handful of smaller seed companies
have been established. However, most small-scale farmers still have little or no access to new
varieties – particularly for open, self-pollinated, and vegetatively propagated crops other than
maize, vegetables and some cash crops like cotton. Many varieties released by national
authorities are rarely multiplied for commercial distribution.
Several factors help explain the limited development of regional seed markets. One
explanation is that high market transaction costs raises the price of seed to unacceptable
levels in the rural market and leads companies to concentrate on a few well established seed
crops that they know farmers will buy (e.g. maize, vegetables, and cash crops). These costs
are reinforced by the high overheads of larger seed companies – including the costs of
maintaining crop breeding programs.
A second explanation is that markets are distorted by the frequent, subsidized distribution of
seed through relief programs. If farmers are able to obtain new varieties through relief
programs, they will not pursue these through the retail market. Further, once a farmer gains
access to a new open- or self-pollinated variety through relief distribution, she will simply
continue to obtain seed from her own crop harvests.
A third explanation suggests that national seed markets are too small to support significant
commercial investment – especially for most secondary crops. This has led to calls for the
harmonization of seed laws and regulations, creating a regional market large enough to
spread marketing risks and promote scale economies.
The combined set of issues ultimately raises questions about the coordination of public and
private sector investments in variety development and dissemination. It takes time to build
seed retail trade systems that farmers can trust and where quality is rewarded by customer
loyalty and competition stimulates innovation leading to a stream of improved varieties that
are needed to sustain productivity increases over time. In the near term, public investments
are needed to speed access to the best available new crop varieties while strengthening
commercial seed sectors. Over time, the public sector will play more of a facilitating role
supporting private investment.

r.jones@cgiar.org
                                                                                             87


47. Genetic diversity among traditional Ethiopian highland maize accessions assessed by
                        SSR markers and morphological traits

              Yoseph Beyene, Anna-Maria Oberholster, Alexander A. Myburg

  Department of Genetics, Forestry and Agricultural Biotechnology Institute, University of
                           Pretoria, 0002 Pretoria, South Africa

Over the past three centuries, maize varieties grown by local farmers have become adapted to
complex environmental conditions in the highlands of Ethiopia. Unfortunately, most of these
traditional varieties are low-yielding. We analyzed 62 traditional Ethiopian highland maize
accessions using 20 simple sequence repeat (SSR) markers and 15 morphological traits in
order to assess genetic diversity and relationships among these accessions and to assess the
correlation between phenotypic and genetic distances in this germplasm. The accessions
varied significantly for all of the measured morphological traits with a marked gradient along
a North to South axis. The average number of SSR alleles per locus was 4.9. Pair-wise
genetic dissimilarity coefficients ranged from 0.27 to 0.63 with a mean of 0.49. Ward
minimum variance cluster analysis showed that accessions collected from the drier Northern
agroecology were distinct from the Western and Southern agroecologies. However, there was
no genetic differentiation between the Western and Southern accessions, suggesting extensive
gene flow, possibly through seed exchange, between these regions. The relationship between
morphological and SSR-based distances was significant and positive (r=0.43, p=0.001). The
high amount of genetic diversity observed in this set of accessions suggested ample
opportunity for the development of improved varieties for the different highland
agroecologies of Ethiopia. However, from a conservation perspective, accessions will need to
be collected and preserved from all of the agroecologies to effectively capture the genetic
variation in the traditional highland maize varieties of Ethiopia.

yoseph.beyene@fabi.up.ac.za
                                                                                               88


       48. Phylogeography of Eucalyptus urophylla based on the chloroplast JLA region

Kitt G. Payn1,2,3, Frank M. Maleka1, Bernard J.H. Janse2, William S. Dvorak3 and Alexander
                                       A. Myburg1,2
 1
     Forest Molecular Genetics Programme, Department of Genetics, Forestry and Agricultural
        Biotechnology Institute (FABI), University of Pretoria, Pretoria, 0002, South Africa
      2
        Mondi Business Paper South Africa, P.O. Box 31024, Merebank, Durban, 4059, South
                                             Africa
     3
       CAMCORE, North Carolina State University, P.O. Box 7626, Raleigh, NC 27695, USA

Eucalyptus urophylla (S.T. Blake) is a fast-growing tropical forest tree species that is often
used in hybrid combinations in South Africa and other eucalypt growing countries. The
natural distribution of E. urophylla is limited to a series of disjunct populations located on
seven islands of the Sunda archipelago in eastern Indonesia. The conservation status of many
of the native provenances range from Vulnerable to Critically endangered. Proficient
management of the natural genetic resources of this tree species with respect to conservation
and breeding strategies requires an operational knowledge of the nature and distribution of
the genetic variation across its native range. Present patterns of genetic variation within the
species have been influenced by evolutionary factors including migration, selection, mutation
and genetic drift. Of particular interest in this study, are the historical migration/colonization
routes of E. urophylla among the aforementioned islands. The chloroplast genome is
maternally inherited in eucalypts and therefore provides a useful marker of seed dispersal
patterns. We have sequenced the hypervariable JLA region of the chloroplast genome of a
species wide sample (n = 160) of E. urophylla. This molecular data is allowing us to study
the relationship between the phylogeny of intraspecific sequence variants (chloroplast
haplotypes) and their geographical distribution across the seven islands of the Sunda
archipelago.

kitt.payn@fabi.up.ac.za
                                                                                             89


       49. Fingerprinting SA cactus pear (Opuntia spp) germplasm using AFLPs

                  Mashope, B.K., Herselman, L., and Labuschagne, M. T.

   University of the Free State, Department of Plant Sciences, P.O. Box 339 Bloemfontein
                                     9300, South Africa

Cactus pear (Opuntia spp.) is increasingly being utilised as an alternative crop in South
Africa for fodder and fruit. Breeding efforts to increase productivity and fruit quality require
cultivar specific information that is currently not available. Previously plant breeders based
their selection of individuals on phenotypic (descriptors) among individuals to develop
distinctive cultivars. Currently molecular genotyping techniques are used in combination
with descriptors, or independently for measuring genetic variation within breeding stocks.
Genotypic characterisation of cactus pear using biotechnology has however been hampered
by the difficulty in extracting gDNA due to the presence of polysaccharide-rich mucilage,
and other secondary metabolites. The objective of this study was to investigate the genetic
diversity within a South African cactus pear germplasm using amplified fragment length
polymorphisms (AFLPs). Thirty eight accessions of cactus pear were analysed using nine
AFLP primer combinations. gDNA was extracted from freeze-dried cladode sections using
the CTAB method. The extracted gDNA was readily fingerprinted using AFLPs. The nine
primer combinations used generated 346 fragments in total, 168 of which were polymorphic.
Genetic similarity estimates were computed and the unweighted pair group method of
arithmetic averages (UPGMA) used for cluster analysis. Analysis of AFLP fingerprinting
data has revealed this as a useful technique to distinguish between the accessions currently
cultivated in South Africa. It is hoped that this information will assist farmers and breeders
by providing a rapid and reliable means of identifying cultivars.

mashopbk.sci@mail.uovs.ac.za
                                                                                           90


  50. Characterization of vernonia (Vernonia galamensis var. ethiopica) as alternative
                        industrial oil crop in Limpopo Province

                       Shimelis Hussein1, P. Mashela1 and A. Hugo2
 1
   University of Limpopo, School of Agricultural and Environmental Sciences, Private Bag
                            X1106, Sovenga 0727, South Africa
  2
    Department of Microbial, Biochemical and Food Biotechnology, University of the Free
                   State, P.O. Box 339, Bloemfontein 9300, South Africa

Venonia (Vernonia galamensis) is a potential new industrial oil seed crop as source of natural
epoxy fatty acids. The oil content and fatty acid profiles of 36 diverse accessions of V.
galamensis variety ethiopica were analyzed to select potential lines for domestication at
Limpopo Province. Accessions showed oil content varying from 22–29% and varying ranges
of vernolic acid (73- 77%), linoleic acid (12-14%), oleic acid (3.5-5.5%), palmitic acid (2.4-
2.9%) and stearic acid (2.3-2.8%). Accessions collected from southern Ethiopia had higher
oil content than those from eastern Ethiopia. Five promising lines, with the highest contents
of oil (28-29%) and vernolic acid (75-77%) were identified as best parents for improving the
quantity of oil and vernolic acid.

shimelis@ul.ac.za
                                                                                         91


 51. Apple cultivar and rootstock development for the South African Second Economy:
                 Enhancing economic and environmental sustainability

                                  Iwan F. Labuschagne

        ARC Infruitec – Nietvoorbij, Private Bag X5013, Stellenbosch, South Africa


Various apple selections and rootstock genotypes with the ability to initiate, develop and
sustain apple production in the South African Second Economy were identified from the
ARC Infruitec – Nietvoorbij apple breeding programme during the past seven years. These
selections were developed from a programme that include breeding goals aimed towards the
diversification of commercial apple production in South Africa i.e., to include developing
farmers and home growers and to increase fruit consumption, nutrition and food safety. The
most important attributes of these selections, include precociousness, high yield potential,
low chilling requirements and resistance against diseases. Some selections are resistant
against apple scab (Venturia inaequalis) and some are derived from precocious families. All
selections were harvested before or during the picking time of ‘Royal Gala’, the earliest
commercial cultivar in South Africa, indicating low chilling requirements. Rootstock
selections include genotypes with resistance against woolly apple aphid (Eriosoma
lanigerum), crown and collar rot (Phytophthora cactorum) and apple mildew (Podosphaera
leucotrica).


labuschagnei@arc.agric.za
                                                                                             92


  52. Comparison of morphological and AFLP genetic diversity analyses of Malawian
                               cassava germplasm

Ibrahim R. M. Benesi1, Maryke T. Labuschagne1, Liezel Herselman1 and Nzola M. Mahungu2
  1.
   Department of Plant Sciences: Plant Breeding, P.O. Box 339, Bloemfontein, South Africa.
                                            9300
                         2.
                           SARRNET, P.O. Box 30258, Lilongwe 3

Commercialisation of cassava coupled with biotic and abiotic stresses lead to genetic erosion.
Information on levels and patterns of genetic diversity is valuable for efficient management and
utilisation of germplasm in breeding programmes to meet the ever-changing needs of growers
and consumers in the face of changing and unpredictable environmental challenges. Therefore,
cassava germplasm was characterised using morphological and AFLP markers. Twenty eight
accessions were characterised using morphological and AFLP methods. Morphological
characterisation was done at Chitedze research satation, Malawi. DNA extraction was done in
Malawi and the rest of AFLP analysis was done at the University of the Free State, South
Africa. AFLP characterisation using cluster analysis showed a narrow range of Dice
similarity coeffient but uniquely differentiated all analysed accessions. Morphological cluster
analysis indicated a wide range of Dice similarity coefficient but could not uniquely
differentiate all characterised accessions. Significant correlation between morphological and
AFLP similarity matrices was observed. Congruencies between morphological and AFLP
dendrograms are also discussed. AFLP analysis confirmed morphological characterisation.
Hence, a combination of AFLP and morphological methods should be planned in such a way
that morphological characterisation using mainly salient traits is conducted first to reduce the
bulk of the germplasm followed by AFLP analysis for morphologically similar accessions,
samples from different morphological heterotic groups and representatives of introductions.

ibenesi@malawi.net
                                                                                            93


 53. Sweetpotato (Ipomea batatas (L) Lam) cultivar mixture by farmers in Malawi: the
                       basis for longer household food security

             Felistas P. Chipungu1, Aggrey J.D. Ambali1 and Nzola M. Mahungu2
  1
      Biology Department,, Chancellor College, University of Malawi, P O Box 280, Zomba,
                                           Malawi
                    2
                      IITA/SARRNET, P O Box 30258, Lilongwe3, Malawi

In Malawi, sweetpotato (Ipomea batatas) is the second important root crop after cassava
(Manihot esculenta Crantz) and the most spread crop grown throughout the country for food
and cash. At policy level, sweetpotato production is being encouraged as it is an important
food security crop in times of maize failure and periods in between maize harvests.
Documentation of indigenous knowledge of traditional varieties of crops is a valuable
complement in the collection, maintenance, and evaluation of diversity representatives as
working collections with purposes of improving traits. In order to collect and document the
amount of cultivar diversity in Malawi and the traditional knowledge associated with the
crop, a survey was conducted in 2003 in seven districts of the country, namely Chitipa,
Karonga, Mzimba, Nsanje, Chikwawa, Mulanje and Phalombe In total, 268 landraces were
collected. These accessions along with introductions were subjected to morphological
characterisation at Bvumbwe Research Station where pre-selection representatives from the
morphological studies were subjected to molecular and nutritional studies at Chancellor
College, the University of Malawi. There was a marked difference in the maintenance of a
variety of cultivars by gender, age, income group and proximity and access to urban centers.
Farmers with low incomes, women and older people of above 50 years grow the most varied
sweetpotato cultivars with the main purpose of reducing risks of food insecurity. Sweetpotato
cultivar diversity in terms of maturity period, shelf life after harvest and resistance to
sweetpotato weevil and mixtures enables household food security over longer periods and
farmers to sustain production to make use of different agro-ecological zones that significantly
vary in terms of soils, topography, altitude, slope, water and fertility. However, in some
areas of study areas which have direct access to urban markets, farmers concentrate on one
high yielding and early maturing cultivar, Kenya whose roots are meant for sale.

felichipungu@yahoo.com
                                                                                             94

                                       POSTERS

P1. Differential response of fourteen sunflower inbred lines to a rust (Puccinia helianthi)
bulk isolate

                        G.Chigeza, T. Debeila, and M.M. Liebenberg

           ARC-Grain Crops Institute, P. Bag X1251, Potchefstroom, South Africa

Sunflower rust, caused by Puccinia helianthi Schw, is becoming a major disease in South
Africa. Research and simulation studies on the effect of rust infection have shown that
reduction in oil yield can be as high as 85% if plants are infested at the vegetative stage. The
objective of this study was to identify potential donors for rust resistance by screening inbred
lines using a rust bulk isolate. Fourteen sunflower inbred lines with resistance to different
races of rust were used. Two separate plantings were done in pots in the greenhouse and each
inbred line had 5-7 plants. Plants were inoculated by spraying with a suspension of P.
helianthi urediospores contained in tap water. Inoculated plants were incubated in dew
chamber for 16-18 hrs and then removed to the green house. Rating on each plant was done
two weeks after inoculation using a score of 1-5, where 1= highly resistant, 2 to 3 =
intermediate resistance, and 4 to 5 = susceptible. Analysis of variance (ANOVA) with
unequal replication was then done for each planting and for the combined data. The disease
ratings varied from 1 to 5. The results were highly significant at 0.01%. Two inbred lines,
IGE SV3 and HAR 2, were the most resistant. HAR 2 is known to be resistant to rust race 5
and the results may indicate that the bulk isolate used is mainly composed of rust race 5. The
correlation of the ratings in the first and second plantings was highly significant (R= 0.82)
indicating high degree of repeatability of the inoculation technique in minimizing escapes.
Further evaluation of the lines will be done with race isolates purified from the bulk isolate
and other field collections to determine which are the prevalent rust races in South Africa.
The pure isolates will then be used in screening rust resistant sunflower lines and varieties
adaptable to the South African conditions. This will increase the efficiency of identifying
donor parents for rust resistance.

chigezag@arc.agric.za
                                                                                             95


P2. Effect of different environments on oil composition in high and mid-oleic sunflower
                                       varieties

                       R. Coetzee1 , A. Hugo2 and M.T. Labuschagne1
  1
  Department of Plant Sciences, University of the Free State, P.O. Box 339, Bloemfontein
                                   9300, South Africa
  2
    Department of Microbial, Biochemical and Food Biotechnology, University of the Free
                  State, P.O. Box 339, Bloemfontein 9300, South Africa

Dietary recommendations favouring high monounsaturates, low saturates and stable
alternatives to hydrogenated oils spurred increased interest in high oleic sunflower oil. Recent
research has lead to the development of high oleic sunflower varieties with oil that may
approach or exceed a 80% oleic acid content. Sunflower oil with high oleic acid content is
less susceptible to oxidative changes during refining, storage and frying compared to
conventional oil. In south Africa, genotype by environment interactions complicate effective
identification of superior sunflower genotypes, leading to variation of relative cultivar yields
across testing environments. Temperature is the major environmental factor affecting oil
content and relative levels of fatty acids in sunflower oil. The objective of this research was
to study oil content and fatty acid composition in high and mid-oleic sunflower varieties
under six different environments. Fatty acid analysis was done using capillary gas
chromatography. Correlations between the major fatty acids were determined as well as the
stability of these fatty acids in different environments. Variability was detected between
different varieties and different environments for oil content and fatty acid composition.
Correlations were detected between some of the major fatty acids.

coetzeer.sci@mail.uovs.ac.za
                                                                                         96


  P3. Conventional breeding of high quality, seedless, Mandarins in the Eastern Cape

                         N. Combrink, Z. Bijzet, and K. Hannweg.

                          ARC -ITSC, PO Box 25, ADDO, 6105

The rapid expansion of the world citrus industry over the past decade has lead to congestion
of the overseas markets. There is a demand for new improved high quality mandarins,
preference given to seedless cultivars. The ARC-ITSC Citrus Breeding program at the Addo
Research Station situated at Addo in the Eastern Cape is focused on breeding these new
improved high quality, seedless, mandarins. This is achieved by combining conventional
breeding and biotechnological techniques. Controlled crosses are made by hand pollinations,
by using a tetraploid female parent and a diploid male parent a triploid embryo is obtained.
The biotechnological technique of embryo rescue is used to rescue this immature triploid
embryo which is then cultured in-vitro to produce a mature triploid plant.

combrinkn@arc.agric.za
                                                                                           97


    P4. Comparison of AFLP and SSR markers for coffee (Coffea arabica L.) genetic
                                diversity analysis

                    Y. Dessalegn, L. Herselman and M.T. Labuschagne

   Department of Plant Sciences, University of the Free State, PO Box 339, Bloemfontein,
                                    9330, South Africa

Coffee belongs to the family Rubiaceae and to the genus Coffea. Although the genus Coffea
is diverse and reported to comprise about 100 species, only two species namely arabica
(Coffea arabica L.) and robusta (Coffea canephora Pierre) are under commercial cultivation.
Arabica coffee’s centre of origin and diversity is in Ethiopia. Coffee is the world’s most
popular non-alcoholic beverage and the second most important commodity in global trade
rated after petroleum products. It is exported in various forms to more than 165 countries and
generates on average US $ 9.7 billion annually. Knowledge of genetic diversity within and
among genotypes of any crop is fundamental to estimate the potential of genetic gain in a
breeding programme and for effective conservation of available genetic resources. This study
was conducted to determine the genetic relationships of 28 C. arabica genotypes collected
from the northwestern and southwestern parts of Ethiopia and compare the efficiency of 10
AFLP primer combinations and six SSR primer pairs in detecting genetic variation among
arabica coffee genotypes and estimate the genetic similarity of coffee genotypes using
combined data of AFLP and SSR analyses. Jaccard similarity coefficients were calculated
and dendrograms constructed using UPGMA method of cluster analysis with NTSYS-pc
software package. AFLP and SSR markers were positively and significantly correlated
(0.217) in estimating genetic similarity among coffee genotypes. The average genetic
similarity coefficient calculated using SSR markers was much lower (0.560 with a range of
0.286-1.000) compared to AFLP markers (0.915 with a range of 0.860-0.982) indicating the
higher information content generated by SSR markers. AFLP markers uniquely distinguished
all coffee genotypes, while SSR markers distinguished 64.3% of the genotypes. The two
dendrograms constructed using either AFLP or SSR analyses data were different. On the
other hand, dendrograms constructed using either AFLP data or combined data of AFLP and
SSR analyses, were similar. Therefore, AFLP markers were more efficient compared to SSR
markers for characterisation of evaluated coffee genotypes.

HerselmanL.sci@mail.uovs.ac.za
                                                                                            98


      P5. Association of bean caffeine content with cup quality and green bean physical
                          characteristics in coffee (Coffea arabica L.)

       Yigzaw Dessalegn1, Maryke Labuschagne1, Gary Osthoff2 and Liezel Herselman1
  1
   Department of Plant Sciences, University of the Free State, P.O.Box 339, Bloemfontein
                                   9300, South Africa
2
  Department of Microbiology, Biochemistry and Food Science, University of the Free State,
                     P.O. Box 339, Bloemfontein 9300, South Africa

Although man started coffee consumption for its stimulating effect, the demand for
decaffeinated coffee is increasing and accounts 10% of the total amount of coffee consumed
in the world. The objectives of this study were to assess caffeine content variability among 42
arabica coffee genotypes from Ethiopia and the associations of caffeine content with cup
quality and green bean physical characteristics. Green bean caffeine content was measured
with HPLC while cup quality was determined by professional coffee tasters. Considerable
variation for caffeine content was observed, ranging from 9.1 to 13.2g kg-1 on dry mass basis
(dmb). Six genotypes, AD0291, AD0591, AD2491, AD2691, AD2791, and AD2891 had a
caffeine content of less than 10.0g kg-1 and may serve as a source of desirable genes for
variety development with relatively low caffeine content. Caffeine content showed negative
and statistically significant associations with cup quality attributes. Therefore, simultaneous
selection both for low caffeine content and better cup quality is possible. Correlations
between caffeine content and green bean physical characteristics were negative but non-
significant, and will not be useful for caffeine content selection in arabica coffee.

labuscm.sci@mail.uovs.ac.za
                                                                                          99


   P6. Identification of root-knot nematode resistance in commercial maize material

                         H. Fourie, A.H. Mc Donald & L. Ngobeni

    ARC – Grain Crops Institute, Private Bag X1251, Potchefstroom, 2520, South Africa

Parasitism by root-knot nematodes (Meloidogyne spp.) is of economic importance in local
maize production. The demand for the identification of resistant maize germplasm is
increasing since chemical control is seldom cost-effective, especially under dry land
production conditions. Local, commercial and small-scale, as well as foreign maize
genotypes were initially evaluated for resistance to M. javanica in greenhouse trials, after
which 22 genotypes were screened for resistance to a mixed population of M. javanica & M.
incognita race 2 in a uniformly infested field. These 22 genotypes also included crosses
between resistant maize lines with inbred lines of appropriate heterotic groupings. In the
greenhouse trial foreign lines were identified with significantly less eggs per root system,
eggs per 50g roots, eggs per g root and lower RF-values than those of a susceptible standard,
which maintained the highest number of M. javanica eggs and larvae and had the highest RF-
value. Identification of molecular markers associated with the resistance trait will
consequently be conducted to facilitate marker-assisted selection to expedite breeding of
root-knot nematode resistant, commercial maize hybrids and varieties.

FourieHD@arc.agric.za
                                                                                           100


                   P7. Using in vitro technology for citrus cultivar improvement
              1
                  Karin Hannweg, 1Zelda Bijzet, 2Nikki Combrink and 1Gerrit Visser
 1
     ARC- Institute for Tropical and Subtropical Crops, Nelspruit, Mpumalanaga, South Africa
                2
                  ARC-Addo Research Station, Addo, Eastern Cape, South Africa

In vitro technologies form an important part of any conventional breeding programme. In
order to improve sales on international markets, it is extremely important for growers to
provide high quality fruit. Furthermore placing fruit on the market very early or late in the
season also ensures that the maximum price is obtained before the market becomes saturated.
Since the genetic base of the sweet orange group is extremely small and breeding difficulties
owing to pollen sterility make it almost impossible to breed improved characteristics into
these cultivars, other methods of cultivar improvement need to be addressed. An important
source of genetic variation is provided by spontaneous mutation in the field in the form of
chimeric fruit. By using in vitro ovule rescue from these chimeric fruit showing excellent
characteristics including: rind and/or flesh colour, rind and/or flesh texture, disease and/or
pest resistance, early or late maturation, appearing as sectors on the fruit, this genetic
variation may be captured in such important cultivars as Navels and Valencias.

In vitro embryo rescue is a technique which allows the rescue of novel genetic material from
controlled tetraploid : diploid crosses. This genetic variation would ordinarily be lost as a
result of embryo abortion 4 months subsequent to the controlled pollinations. This
experiment is focussed on rescuing triploid (i.e. embryos which produce seedless fruit)
embryos, in vitro, of several controlled crosses carried out by the conventional breeding team.
Plantlets obtained from the embryo rescue process are then placed into the evaluation
programme.

The posters describe the potential impacts of these technologies on citrus cultivar
improvement in South Africa.

karin@arc.agric.za
                                                                                            101


                P8. New sweet potato cultivars for resource-poor farmers

          Sunette M Laurie, M.D Magoro, M.M. Mtileni, and A.A. Van den Berg

   Agricultural Research Council – Roodeplaat Vegetable and Ornamental Plan Institute,
                              Pretoria, 0001, South Africa

Some of the main restricting factors in sweet potato production by resource-poor farmers are
low yields and yield instability as a result of the use of old landraces. The availability of
improved cultivars would increase the sustainability and productivity of sweet potato
production, thereby contributing to food security. A very important trait for these consumers,
is a sweet and dry taste opposed to the watery texture of commercial cultivars.
Sweet potato breeding at ARC-Roodeplaat focused on the development of cultivars with
good yield, good storage quality combined with sweet and dry taste. The breeding procedure
entailed importation of novel germplasm, polycrossing (9000 seeds), stringent selection at the
2000-3000 seedlings in the seedling nursery, evaluation of 100-150 entries in the preliminary
yield trial, 20-25 in the intermediate yield trial and 15 in the advanced yield trial per annum.
Advanced genotypes were evaluated further in off-station trials in target production areas
during 2000/01 to 2004/05 in order to determine adaptability and acceptability, and then
release cultivars for production. Farmer participatory selection was a very important aspect of
the process. Recommendations were made based on yield as well as taste acceptability.
The result was the registration of plant breeder’s rights on seven new cream-fleshed cultivars
(Ndou, Monate, Mokone, Letlhabula, Phala, Amasi and Mamphenyane), one orange-cream
cultivar (Serolane) and one orange-fleshed cultivar (Khano) developed at ARC-Roodeplaat.
Additionally, imported orange-fleshed cultivars Excel, W-119 and Resisto from the USA and
A15 from University of KZN, were recommended for production. Orange-fleshed sweet
potato contains high levels of pro-vitamin A and can contribute significantly to alleviation of
vitamin A deficiency.

slaurie@arc.agric.za
                                                                                            102


                  P9. Development of a new Ur-11 marker in dry beans

                  L.A. Madubanya, M.M. Liebenberg and C.M.S. Mienie

     ARC-Grain Crops Institute, Plant Breeding and Biotechnology, Private Bag X1251,
                                   Potchefstroom, 2520

Rust, caused by a fungus Uromyces appendiculatis (Pers.) Unger, has a significant effect on
the yield and quality of beans in many parts of the world. There are over 300 races
(pathotypes) of this fungus recognized. Conventional control methods of bean rust are costly
and environmentally unfriendly. The most economic form of control is the genetic resistance
present in the host plant. Single dominant genes control this resistance, although other reports
have suggested single or multiple dominant and recessive genes being involved. There are at
least 13 resistant genes
(Ur-11 to Ur-13) identified to date, organized in the genome as clusters, since they confer
resistance to multiple races of U. appendiculatis. The gene Ur-11 is present in the
Guatemalan black bean PI 181996 (Middle-American origin). This cultivar is resistant to all
known rust races in the USA but the Ur-11 gene is epistatic to other less effective resistance
genes, Ur-4 and Ur-5. There is therefore a need to develop markers tightly linked to Ur-11.
Several markers linked to Ur-11 were reported but records of their effective use for Marker
Assisted Selection (MAS) outside the original mapping populations were not found. We
report on the detection of possible marker(s) in the F2 population segregating for the Ur-11
resistant gene using Bulked Segregant Analysis (BSA). DNA was extracted from F2 lines
resulting from a cross between a susceptible cultivar from our local gene pool (Kranskop)
with the resistant PI 181996 of Middle-American origin. Two contrasting bulks, resistant and
susceptible, were generated and screened for informative polymorphisms using amplified
fragment length polymorphism (AFLP) analysis. In total 75 primer combinations were tested,
with Kranskop and PI 181996 serving as controls. Thirty-two primer sets yielded informative
polymorphisms and these were used in the amplification of individual plants. Only 10 primer
sets yielded 12 putative markers that showed co-migration with the resistant gene. Of the 12,
six were linked to the resistant gene in coupling phase while the remainder were in repulsion.
More F2 plants from the segregating population are being amplified with the selected 10
primer sets to verify these putative markers. Once verified, these possible AFLP markers for
Ur-11 will be converted into sequence-characterized amplified regions (SCARs). This will
aid plant breeders in the screening of plants that carry the Ur-11 resistance gene in our local
gene pool.

madubanyal@arc.agric.za
                                                                                              103


P10. Alternative early generation selection methods to predict better malting quality in
                               barley (Hordeum vulgare)

                       A.F. Malan, H.A. Smit, R. Oelofse and M. Ncala

                       Small Grain Institute P/B x29, Bethlehem, 9700

The barley cultivar Clipper was introduced in South Africa in the mid 1970`s. Ever since
then not one released cultivar was able to exceed Clippers` performance with regard to
malting and brewing quality. Clipper currently contributes up to 60% of the annual crop in
the Southern Cape barley production area. The South African maltsters and brewers have
shifted their preference from European and Southern Hemisphere to North American malting
and brewing types. This adds to the challenge of barley plant breeders to change their barley
quality selection criteria in the shortest time possible. Up to now quality evaluation was only
possible in the F5 generation when enough seed is available. The shift in South African
malting and brewing ideotipes accentuate the need for new quality selection methods in early
generations of a breeding programme.

Barley malt and brewing quality are genetically coded by approximately 559 polygenic
fractions. It is therefore necessary to evaluate alternative selection methods to fully exploit
these genetic potential in early generations. Three quality parameters were evaluated on the
seed of 18000 and 4500 single plants in F2 and F3 generations respectively, namely kernel
plumpness, kernel nitrogen content and germination energy. Results obtained for these
characteristics gave a normal distribution curve, which is typical for polygenic traits. From
the results it is evident that only 0.04% of the F2 population fulfill in the quality requirements
for the three traits measured. However, if the selection procedure for malt and brewing
quality were postponed until the F3 generation, only 0.007% of the population would meet
the specifications.

malane@arc.agric.za
                                                                                              104


                   P11. Quality Protein Maize (QPM): facts and figures

                                   Kingstone Mashingaidze

                        ARC-GCI, P/Bag X1251, Potchefstroom, 2520

Maize (Zea mays L.) is the staple food crop and mainstay of diets of smallholder farmers in
South Africa. The nutritive value of normal maize is rather poor because of a marked
deficiency of its protein in two essential amino acids (tryptophan and lysine), the presence of
excess amount of leucine and low availability of niacin. The low availability of niacin is
further worsened by a marked deficiency of tryptophan, its precursor. In addition, the high
amount of leucine is known to inhibit the conversion of tryptophan to niacin. As a
consequence, pellagra, the niacin deficiency disease, is associated with diets almost solely
based on maize.
         The low nutritive value of maize can be corrected in genetically improved Quality
Protein Maize (QPM). The opaque-2 gene confers higher levels of lysine and tryptophan to
maize grain. QPM protein contains 60 – 100% more lysine and tryptophan, and 38% less
leucine than normal maize. The remarkable increase in tryptophan and parallel decrease in
leucine favours the synthesis of niacin, resulting in improved niacin status of those who use
maize as a main component of the diet. Due to these characteristics:
the biological value, or the amount of nitrogen that is retained in the body, is about 80% for
QPM, compared to 40 – 57% for normal maize and 86% for eggs.
Protein of normal maize has about 40%, whereas QPM has 90% the biological value of milk.
The net protein utilization, which is the product of digestibility and biological value, is about
40% for normal maize and 65% for QPM.
If compared to FAO/WHO standard protein, the QPM chemical score ranges from 90 to
100%, while that of normal maize is 51%.
         There is no doubt that QPM is nutritionally superior to normal maize. Individual case
studies have shown the remarkable recovery of children suffering from severe protein
malnutrition, when QPM was substituted for normal maize in their diet. In infant feeding
trials in Ghana and Ethiopia it was reported that (i) infant growth, stunting, duration of illness
and mortality, were all improved through the substitution of QPM for normal maize, and (ii)
wherever maize is an important and/or dominant weaning food, QPM has better growth
enhancing capabilities than does normal maize and results in far less stunting. QPM has also
shown even more dramatic results in experiments with pigs and chickens.
         The value of QPM in fighting hunger and malnutrition has advantages over
approaches using non-conventional foods that require changes in diet habits. QPM is not a
new crop. QPM provides an inexpensive high quality protein and energy source for
combating malnutrition, at no additional cost to farmers. QPM has the promise of improving
the nutritional status of all vulnerable groups whose main staple is maize and who cannot
afford enough protein rich foods to supplement it. QPM may be also valuable in feeding
programmes, particularly for school Feeding Programmes and home-based care programmes
for people living with HIV/AIDS.
         There is, therefore, a strong case for the replacement of normal maize varieties with
QPM in communities that depend on maize as an important source of calories and protein in
their diets.

MashingaidzeK@arc.agric.za
                                                                                           105


         P12. Inhibitory activities of two apple Polygalacturonase Inhibiting Proteins
                                           (MdPGIPs)

                      L.B.T Matsaunyane1, D. K. Berger2 and D. Oelofse1
    1
    Agricultural Research Council Roodeplaat, Private Bag X293, Pretoria, South Africa, 0001
2
    Department of Botany, Forestry and Agricultural Biotechnology Institute (FABI), University
                            of Pretoria, Pretoria, South Africa, 0001

Plant diseases have always resulted in significant crop losses, and traditionally, a number of
methods have been used to minimise or eliminate the harmful effects of plant disease. These
include conventional breeding for the production of disease resistant cultivars, and the
application of fungicides. There are, however, several disadvantages associated with these
strategies. One of the newer alternatives used to combat plant disease is the engineering for
fungal resistance in transgenic crops. The discovery of plant antifungal proteins and the
isolation of their encoding genes assist in the engineering of plants with increased resistance
towards certain fungal pathogens.

During pathogenesis, fungal polygalacturonases (PGs) degrade the cell wall pectin, allowing
the pathogen to penetrate the plant. Polygalacturonase-inhibiting proteins (PGIPs) are cell
wall-associated proteins that inhibit PG activities by forming specific, high-affinity
complexes with them. PGIPs from a single plant source have shown different inhibitory
activities against PGs from the same pathogen. Phaseolus vulgaris PGIP1 (PvPGIP1) is
unable to inhibit Fusarium verticilliodes PGs, whereas P. vulgaris PGIP2 (PvPGIP2) extracts
interacted strongly with these PGs.

Apples (Malus domestica) contribute a relatively large percentage to the total agricultural
production in South Africa. Several factors influence apple production, including damage and
diseases. Diseases caused by phytopathogenic fungi have the most influence, particularly
during fruit storage. These findings prompted a study of the inhibitory activities of M.
domestica PGIPs (MdPGIPs) against several fungal PGs.

Two pgip genes have been isolated from M. domestica cv Granny Smith plants (termed
Mdpgip1 and Mdpgip2) at ARC-Roodeplaat. Both the Mdpgip1 and Mdpgip2 genes were
expressed, individually, under control of the constitutive CaMV dual 35S promoter, in
transgenic Nicotiana tabacum. MdPGIP extracts from these plants inhibited Botrytis cinerea,
Botryosphaeria obtusa, Diaporthe ambigua and Verticillium dahliae PGs. These results
indicated that the isolated Mdpgip genes encode active proteins. Further inhibition studies
will be performed to find a fungal PG which one MdPGIP inhibits, but the other MdPGIP
does not. In conclusion, the gene encoding an effective PG inhibitor can be used to produce
transgenic plants exhibiting a degree of resistance to certain fungal pathogens.

lmatsaunyane@arc.agric.za
                                                                                             106


 P13. Halo blight resistance in host differential cultivar ZAA 12 is conditioned by three
                                           genes
                              1
                                  Phillip Miklas and 2Deidré Fourie
   1
       USDA-ARS, 24106 N. Bunn Road, Prosser, WA, 99350 (pmiklas@pars.ars.usda.gov);
              2
                ARC-Grain Crops Institute, Potchefstroom, 2520, South Africa

Halo blight (Pseudomonas syringae pv. phaseoliciola) is a serious seed-borne bacterial
disease that limits dry bean (Phaseolus vulgaris L.) production worldwide. Monogenic
resistance is effective against most races of the pathogen. ZAA 12 (A43) is the most resistant
host differential cultivar conditioning resistance to seven of the nine differential races of the
pathogen. Our objective was to study the inheritance of resistance in ZAA 12 to Races 2, 3, 4,
5, 7, 8, and 9. Seventy-nine F6:7 RILs from ZAA 12/Canadian Wonder were infected with
seven races in separate inoculations. Results indicate presence of Pse-3 gene conditioning
hypersensitive resistance to Races 3 and 4; Pse-4 gene for resistance to Race 5; and an
unknown gene governing resistance to Races 2, 7, 8, and 9. Allelism tests are underway with
other host differential cultivars to confirm independence of this locus from Pse-1 gene.

FourieD@arc.agric.za
                                                                                        107


   P14. Host suitability of dry bean germplasm to the root-knot nematode species M.
                                        javanica

                                  H Muedi and H. Fourie

  Agricultural Research Council-Grain Crops Institute, Private Bag X1251, Potchefstroom,
                                   2520, South Africa

Root-knot nematodes are economically important pathogens of dry bean, which is a life-
sustaining crop for small-scale farmers and an economically important crop for commercial
producers in South Africa. Since no local dry bean genotype exhibited resistance to M.
javanica and M. incognita race 2, respectively, foreign dry bean breeding lines with known
resistance to root-knot nematodes were obtained for evaluation in this regard. Subsequently
these foreign dry bean genotypes, together with local, root-knot nematode susceptible
standards were evaluated for host suitability to M. javanica in a greenhouse experiment. Dry
bean seedlings were artificially inoculated with approximately 10 000 M. javanica eggs and
second-stage juveniles (J2) twelve days after planting and nematode assessments were done
56 days later. Significant differences were obtained between genotypes with regard to all
nematode parameters evaluated. Local dry bean genotypes generally exhibited high ELF-
indices, number of eggs and J2 per root system as well as RF-values. On the other hand,
several foreign genotypes were identified as poor hosts to this root-knot nematode species.
These foreign dry bean breeding lines will also be evaluated for host suitability to M.
incognita race 2 and will subsequently be incorporated into the local dry bean breeding
programme. Breeding for root-knot nematode resistant dry bean genotypes could offer long-
term and sustainable solutions to dry bean producers for effective management of root-knot
nematode infestations.

MuediH@arc.agric.za
                                                                                          108


P15. Purification of apple polygalacturonase inhibiting proteins 1 and 2 from transgenic
       tobacco, and an investigation into their inhibitory activities against fungal
                                   polygalacturonases

  D. Oelofse1, I.A. Dubery2 , R. Meyer2, I. Gazendam1, G. De Lorenzo3 and D. K. Berger4
              1
                Agricultural Research Council (ARC)-Roodeplaat, South Africa
                2
                  Dept. Biochemistry, University of Johannesburg, South Africa
            3
              Dipto di Biologia Vegetale, Universita di Roma “La Sapienza”, Italia
   4
     Dept. Botany, Forestry and Agricultural Biotechnology Institute (FABI), University of
                                    Pretoria, South Africa

Polygalacturonase-inhibiting proteins (PGIPs) are plant cell wall proteins that counteract the
action of fungal polygalacturonases (PGs) and reduce fungal disease symptoms by hampering
the invasion process and the release of nutrients necessary for fungal growth. Two PGIPs
(MdPGIP1 and MdPGIP2) of Malus x domestica were separately expressed in transgenic
tobacco plants and purified to apparent homogeneity by means of anion and cation exchange
chromatography to investigate their inhibitory activity. PGs from the lupin pathogen
Colletotrichum lupini, two apple pathogens, Botryosphaeria obtusa and Diaporthe ambigua,
two maize pathogens Fusarium verticillioides and Stenocarpella maydis, the onion pathogen
Botrytis cinerea and Aspergillus niger. MdPGIP1 and MdPGIP2 were tested. MdPGIP1
appeared to be a more effective inhibitor of PGs from the apple pathogens, since twice as
much MdPGIP2 (20 ng) was required for 50% inhibition of a crude PG preparation of C.
lupini in an agarose diffusion assay. Both MdPGIP1 and MdPGIP2 were strong inhibitors of
partially purified PGs from S. maydis and B. cinerea, requiring only 6 ng for 50% PG
inhibition. In contrast, 30 ng of either PGIP was required for 50% inhibition of a purified PG
from C. lupini. Neither of the MdPGIPs was able to inhibit purified PGs from F.
verticillioides or A. niger. We conclude that there are no differences in the spectrum of PGs
inhibited by the two MdPGIPs.

doelofse@arc.agric.za
                                                                                             109



                    P16. Studies on the S-locus and self-incompatibility in apple

          1
              Brendon Okkers, 1Joseph Mafofo, 2Iwan Labuschagné and 1D. Jasper G. Rees
          1
              Department of Biotechnology, University of the Western Cape, Bellville, 7535
  2
      Breeding and Evaluation Division, Agricultural Research Council, Infruitec-Nietvoorbij,
                                       Stellenbosch, 7599

Successful pollination in apples and pears is controlled by a self-incompatibility system,
which is determined by a single multi-allelic S-locus. To determine S-allele genotypes for
some of the cultivars in the ARC’s breeding stock, leaf material was collected and used to
isolate DNA for genotyping experiments. In total 89 samples were collected and genotyped
using 15 universal allele-specific primers previously published by Broothaerts (2003). S-
alleles that co-amplified with the same primer pairs such as S4; S27a; S27b and S20;
S14/17/21 were further discriminated using restriction digestion and resolving digestion
profiles on agarose gels. All the 89 cultivars were genotyped using this approach. Recent
studies in Prunus and some other species have shown the presence of a polymorphic F-box
gene, closely linked to the S-locus. Therefore in order to study the pollen-S determinant of
gametophytic self-incompatibility in apple and pear, sequence data from previously cloned
and sequenced F-box genes were downloaded and aligned. Highly conserved regions were
chosen, and primers designed for cloning of F-box family genes. These gave a fragment in
the expected 500bp size range. PCR amplicons from reactions on ‘Golden Delicious’ were
cloned and sequenced. The analysis of this gene will increase our understanding of the self-
incompatibility system, and this will be of value both for the breeding program and for the
selection of cross pollinators in commercial orchards.

jmafofo@mail.biotech.uwc.ac.za
                                                                                           110


                     P17. DNA fingerprinting of table grape cultivars

           R. Prins1, C.J. van Heerden2, A.L. Burger3, P. Burger4 and W.A. Smit4
                1
                   CenGen, 78 Fairbairn Street, Worcester, 6850, South Africa
2
  Central DNA Sequencer, Stellenbosch University, P/Bag X1, Matieland, 7602, South Africa
3
  SunBio, Institute Wine Biotechnology, Stellenbosch University, P/Bag X1, Matieland, 7602,
                                         South Africa
      4
        ARC-Infruitec-Nietvoorbij, Private Bag X5026, Stellenbosch, 7599, South Africa

Developmental and environmental factors affect expression of morphological characteristics,
complicating the traditional methods of identifying grapevine (Vitis vinifera L.) cultivars.
DNA-based molecular markers, like Simple Sequence Repeats (SSRs, also known as
microsatellites), are not affected by these factors. SSRs, which are relatively abundant in the
genomes of all higher organisms, have been shown to exhibit sequence variation between
grapevine cultivars. In this study seventeen table grape cultivars, of which eleven are
commercially grown in South Africa, were selected for SSR analysis. In an attempt to limit
costs, seven fluorescently-labelled SSR primer pairs were combined in a single multiplex
PCR reaction. This, in combination with automated analysis of the PCR products on an ABI
3100 genetic analyser, resulted in informative and repeatable data that could be produced in a
relatively short time compared to other DNA-based molecular markers. This study showed
that the small set of seven SSR markers could be used to discriminate between the seventeen
table grape cultivars subjected to the analysis. The highly polymorphic nature of the chosen
SSR markers suggests that they will be useful in the typing of more of the commercially
important table grape cultivars. In order to improve transferability across different genetic
analysers, preliminary work to determine the feasibility of developing an allelic ladder as
internal control for the DNA fingerprinting of table grapes, has been done.

cprins@lantic.net
                                                                                             111


P18. Analysis of QTLs affecting dormancy release in apple (Malus x domestica Borkh.)
1
    Maria M. van Dyk, 1M. Callies Selala, 1Zolani Simayi, 1Sonwabo Booi, 1Ramsey Maharaj,
              1
                M. Khashief Soeker, 2Iwan F. Labuschagné. and 1D. Jasper G. Rees.
            1
            Department of Biotechnology, University of the Western Cape, Bellville, 7535
    2
        Breeding and Evaluation Division, Agricultural Research Council, Infruitec-Nietvoorbij,
                                         Stellenbosch, 7599

Winter temperatures in the Western Cape region of South Africa are not cold enough for
normal dormancy release in apple trees during spring. Initial vegetative budbreak has been
shown to be associated with prolonged dormancy symptoms experienced by local farmers.
Marker assisted selection (MAS) of apple cultivars with a lower chilling requirement will be
beneficial for local production and will ensure that South Africa remains one of the biggest
competitors on the global export market. In this study 5 mapping populations have been
analysed for time of initial vegetative budbreak, and these provide the basis for the analysis
of the genetic control of this phenotype. The first step towards MAS is the generation of a
genetic linkage map. We describe the development of 310 new (write out) SSR markers,
using apple and pear sequences, containing di-, tri- and tetranucleotide repeats. The newly
developed markers, together with 193 published markers, are being used for cultivar
identification studies. Polymorphic markers are used to screen the progeny of the five
controlled crosses made in order to study and identify quantitative trait loci (QTL) affecting
dormancy release after winter. The prediction, optimisation, multiplexing and mapping of the
complete set of markers is an ongoing process, and the current status of the maps will be
described. The identification of genetic markers linked to QTLs controlling the time of bud
break will allow the selection of the desirable genotypes in future marker assisted selection in
the apple breeding program.

mvandyk@uwc.ac.za
112

				
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