QTL Mapping of Polyphenol Oxidase _PPO_ Activity and Yellow

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www.thaiagj.org Thai Journal of Agricultural Science 2010, 43(2): 109-118

QTL Mapping of Polyphenol Oxidase (PPO) Activity
and Yellow Alkaline Noodle (YAN) Color Components
in an Australian Hexaploid Wheat Population

A. Sadeque1,* and M.A. Turner2

1
Department of Industry and Investment, Narrabri, NSW, Australia
2
The University of Sydney, Plant Breeding Institute, Cobbitty, Australia

*Corresponding author. Email: abdus.sadeque@industry.nsw.gov.au

Abstract
Polyphenol oxidase (PPO) catalyses the undesirable darkening of wheat products such as Asian
noodles. Genetic variation for PPO activity is present in breadwheat and low PPO activity is a
current target of Australian wheat breeding programs. An improved understanding of the genetic
control of PPO activity in wheat backgrounds that are relevant to yellow alkaline noodle (YAN)
wheat production in northern NSW and QLD would expedite this process. QTL (Quantitative Trait
Loci) mapping of polyphenol oxidase (PPO) activity was investigated in a doubled haploid (DH)
population that was derived from a hybrid between a low PPO line DM5637*B8 and high PPO
Australian wheat variety H45. These trials were conducted at The University of Sydney Plant
Breeding Institute, Narrabri, NSW during the 2005 and 2006 growing seasons. In both seasons
PPO activity was significantly (P<0.001) correlated with changes in brightness (∆L*) and
yellowness (∆b*) of YAN. . QTLs on chromosome 2A (QPPO.dmh45-2AL, Q∆L*.dmh45-2AL,
Q∆b*.dmh45-2AL) were associated with the characters at a high level of significance (P>0.001)
and had LRS values of 59.9, 47.7 and 53.8, respectively. In each case the identified QTL explained
more than 50% of the phenotypic variation of the traits. A highly significant (P<0.001) association
was identified between marker locus wPt-7024 (chromosome 2A), and PPO activity, ∆L* and
∆b*. This information may be applied to increase the efficiency of breeding efforts that aim to
generate improved YAN wheat.

Keywords: polyphenol oxidase (PPO), doubled haploid (DH), QTL mapping, Asian noodle,
YAN, colour

Introduction contributor to browning of noodles, chapattis and
other wheat products (Demeke et al., 2001a).
Wheat flour noodles are an important part of the Polyphenol oxidase (PPO) catalyses the oxidation
diet in many Asian countries. Noodles that are of phenolic compounds to quinones, which undergo
bright and have stable color are preferred by further rearrangement, non enzymatic oxidation,
consumers. Those that are commonly sold fresh are and polymerization to produce dark or brown
sensitive to time dependent darkening which is colored melanin (Whitaker and Lee, 1995; Demeke
undesirable (Baik et al., 1995; Hou and Kruk, 1998; et al., 2001). Due to oxidation, this action causes
Hatcher et al., 1999; Jimenez and Dubcovsky, discoloration of some wheat products including
1999). Oxidative enzymes such as polyphenol Asian noodles (Kruger et al., 1992; Baik et al.,
oxidase (PPO), lipoxygenase (LOX), peroxidase 1995, Fuerst et al., 2006), pasta (Simeone et al.,
and catalase cause time dependant darkening of 2002), pan breads and steam breads (Dexter et al.,
wheat products (Feillet et al., 2000). PPO is a major 1984). Cultivars with low PPO activity are
110 A. Sadeque and M. A. Turner Thai Journal of Agricultural Science

desirable for consumers and food manufacturers. (AUS1408/Sunco), a line with low PPO activity,
The understanding of the influence of PPO activity sprouting tolerance and low blackpoint incidence
on yellow alkaline noodle color is required by (Howes pers. comm.) and Australian cultivar H45
breeders that are developing wheat cultivars that (Ciano67/2*Olympic/3/WW80/3*Anza//Kalyonson
may be used as an input of yellow alkaline noodle a/Bluebird) was used in this study. The doubled
manufacture. The influence of PPO activity on haploid population was developed using the wheat
YAN darkening is known but mapping approaches x maize system (Ahmed pers. comm.; Johnson et
concerned with YAN color and color stability have al., 2005). For the PPO experiments, plants were
rarely been used in Australian wheats. sown in 3m rows and a sub set of 48 lines and
Variation in PPO activity is reported between parents were grown in 2 m x 6 m plots for the
wheat cultivars in different locations (Baik et al., yellow alkaline noodles (YAN) testing. A RCBD
1994; Park et al., 1997). The growing environment design with 2 replications was employed for both
has an influence on PPO activity in wheat (Baik et experiments. Trials were conducted at the Plant
al., 1994; Park 1997) and negatively impacts on the Breeding Institute, Narrabri during the 2005 and
efficiency of selection for low PPO genotypes 2006 growing seasons.
(Demeke et al., 2001). Furthermore, PPO activity is
expressed in maternal tissue and it is difficult to PPO Assay
select for low PPO activity early in the breeding The grain PPO activity was determined by using
cycle using standard substrate assays in segregating tyrosine as substrate for the assay procedure as
material. Because it might has the consequence of described by Bernier and Howes (1994) and
reducing population size. Identification of McCaig et al. (1999) except that the incubation
molecular markers that can select wheat lines with period was increased from 2.5 to 3.5 h and the
low PPO phenotypes will enhance the efficiency of optical density was determined at 450 nm by a
plant breeding programmes that have low PPO Bench Mark Plus-Microplate Spectrophotometer
activity as a target. Loci on the long arm of (BIORAD laboratories 2000, Alfred Nobel Drive,
chromosomes 2AL (Raman et al., 2005; Sun et al., Hercules, CA 94547). Twenty whole seeds for each
2005; Zang et al., 2005; He et al., 2007) and 2DL line were incubated in 0.01 M disodium tyrosine
(Zang et al., 2005; He et al., 2007) in hexaploid solution with 0.2% Tween 20 at 37oC for 3.5 h, and,
wheat have a major genetic effect on PPO activity after leaving the seeds, the absorbance of the
in some genetic populations. Recently, He et al. solution was measured at 450 nm.
(2007) found that a combination of markers
(PPO33/PPO16) for chromosomes 2A and 2D loci Yellow Alkaline Noodles (YAN)
was an efficient and reliable means of selecting low The yellow alkaline noodle evaluation
PPO genotypes. Hence, the present investigation experiments for the crop grown in 2005 and 2006
aimed to identify the associations between PPO were conducted in Western Wheat Quality
activity and yellow alkaline noodle color in a Laboratory (WWQL), Washington State
doubled haploid (DH) population derived from University, USA and the Plant Breeding Institute,
DM5637*B8 (low PPO line) x H45 (high PPO University of Sydney (PBIN), Narrabri, NSW,
line). It also aimed to identify genomic regions that Australia, respectively.
control PPO activity and color stability of YAN and
molecular markers that will be useful to select Flour Milling
wheat lines with high color stability of yellow Grains of the lines were milled using a
alkaline noodles. Quadrumat Junior Mill (Brabender® OHG
DUISBURG, Germany). The flour extraction rate
Materials and Methods was adjusted to 60 percent. Moisture and protein
contents were measured using Near Infrared
Plant Materials Reflectance (NIR; Perten Instruments, Type
A doubled haploid population (n = 187) that was 9100/01, Germany). The samples were conditioned
generated from a hybrid between DM5637*B8 to 15% moisture overnight by adding the
Vol. 43, No.2, 2010 QTL mapping of PPO activity and YAN color in wheat population 111

appropriate amount of water before milling. Flour extraction buffer was prepared with 2% (w/v)
and bran yield was recorded for each sample. CTAB (sigma), 1.4 M NaCl, 0.2% mM EDTA, 100
mM Tris-HCl, (pH 8.0).
Yellow Alkaline Noodle (YAN) Sheet
Preparation DArT Marker Analyses
At WWQL, The noodle sheet was produced as 2.5 µg of restriction quality DNA of each DH
described by Morris et al. (2000). The stock genotype (n=92) and the parents of the population
solution was prepared by mixing 200 g NaCl with DM5637*B8 x H45 was sent to Triticarte, Canberra
50 g of Na2CO3 in 1000 mL of H2O in a volumetric (Australia; http://www.triticarte.com.au) for whole
flask. Five mL of this stock solution containing 1 g genome profiling using Diversity Array Technology.
of NaCl and 0.25 g of Na2CO3 was added to 50 g of The 92 lines that were the subject of this study were
flour in this test. At PBIN, the stock solution was randomly selected from the population. The loci were
prepared according to the method of Mares and scored as present (1) or absent (0) and the locus
Campbell (2001) by mixing 25 g NaCl, 15 g KCO3 designations were as used by Triticarte Pty. Ltd. The
and 10 g Na2CO3 with 875 mL water. The yellow P value reflects how well the two phases (present=1
alkaline noodle sheets were prepared by adding 10 vs absent=0) of the marker are separated in the
g of flour with 3.7 mL of alkaline noodles solution. sample and P is based on ANOVA which is an
estimate of marker quality. Markers with P values 80,
Yellow Alkaline Noodle (YAN) Sheet and Flour 77-80, and 75-77 are termed as extremely reliable,
Color Measurement usually scored, and provide useful information,
Wheat noodle sheet color (CIE L*, a*, b* respectively (Wenzl et al., 2004; Huttner et al., 2006).
denotes lightness which is white-black, red-green,
and yellow-blue scales, respectively) were measured Linkage Map Construction and QTL Mapping
with a chromameter (Model-300, Minolta Camera The initial linkage mapping was performed with
Co., Ltd., Osaka Japan) with a 50 mm diameter Cartablanche software, version 1.5.0(111), Keygene
measuring tube using a white tile background. The Products B.V. Linkage groups were further
average of three readings, that were taken after reassessed and reconstructed with Map Manager
moving the head each time, was employed in (QTXb20). The assignment of markers in linkage
analysis. Four color readings at 0, 2, 24and 48 h (2 groups for individual chromosome relied on the
h reading was not taken at WWQL) were taken. consensus map of the Triticarte Ltd. and ongoing
Noodles sheets were stored in plastic bags at 21oC construction of consensus map for the DArT assay
between color readings. The change in color values (Akbari et al., 2006; Howes pers. comm.; http://
(CIE ∆L*,∆a* and ∆b*) was calculated by www.triticarte.com.au)
subtracting readings at 2, 24 and 48 h from those Interval mapping was performed at P=0.01 for
immediately taken after noodle sheets were made marker-trait association using Map Manager
(time zero). Values from duplicate noodle sheets QTXb20 (Manly et al., 2001). Single marker
were used in all of the analyses. regression and simple interval mapping tools were
used for this purpose. The marker regression function
DNA Isolation (P=0.01) was performed to find single marker loci
Genomic DNA extractions were performed linked with the quantitative data. The analysis was
using a modified CTAB protocol according to carried out for each of the two individual
Doyle et al. (1990). 2.5µg of restriction quality environments/years and for the pooled environments.
DNA of each DH genotype and the parents of the The LRS thresholds in the regression analyses
DM5637*B8 x H45 population were prepared. P<0.01 and P<0.001 were used for declaring
Genomic DNA was extracted from fresh leaf tissue significant and highly significant levels, respectively.
(0.5 to 1g) from a bulk of four plants. Leaves from The Map Manager QTXb20 (Manly et al., 2001) and
three weeks old plants were taken. Leaves that were MapChart version 2.2 (Voorrips 2002) were used to
dry, disease free and free from sticking soil generate QTL map and chromosome map figures for
particles were selected for DNA extraction. CTAB presentation.
112 A. Sadeque and M. A. Turner Thai Journal of Agricultural Science

Results brightness (L*) at 24 hours was clearly influenced
by PPO activity (i.e. lower PPO activity = YAN
PPO Assay brightness) of the genotypes.
The PPO activities determined for each of the
lines were consistent across years and were Noodle Sheet Color Trait b*
significantly correlated (r2=0.38, P<0.001; Figure The ∆b* values were significantly correlated
1). The low PPO parent, DM5637*B8 had (r2=0.53 and 0.67, P<0.001) with PPO activity in
consistently low PPO activities in 2005 and 2006 2005 and 2006 (Figure 3). Results indicate that low
(A450 of 0.229 and 0.274, respectively). The PPO PPO activity is associated with elevated ∆b* in this
activity of H45 was high in both seasons (A450 of population.
0.400 and 0.559, respectively). The PPO activity
(A450) of lines ranged between 0.144 to 0.603 with a
population mean of 0.314 in 2005. In 2006, the A450
ranged from 0.164 to 0.698 and the mean for the y = 0.78x + 0.12
population was 0.367. In both seasons the frequency 0.70 R2 = 0.38
n=189
distributions of PPO activity were near normal (data

PPO activty (A ) Year-2006
0.60

not shown). Transgressive segregation was evident 0.50
in this population. However, sixty nine (37%) lines

450
0.40
in the population possessed a significantly higher
PPO activity than the low PPO parent DM5637*B8 0.30

across seasons (LSD0.05=0.137). 0.20

0.10
Noodle Sheet Color Trait L* 0.10 0.20 0.30 0.40 0.50 0.60

The correlation between PPO activity (A450) and PPO activity (A 450) Year-2005

YAN (∆L* [0-24h]) was significant r2=0.52 and
Figure 1 Relationship between PPO activity of lines
0.78 (P<0.001) in 2005 and 2006, respectively from DM5637*B8 x H45 DH population in 2005 and
(Figure 2). Results of both years revealed that YAN 2006 seasons.

(a) Year 2005 (b) Year 2006
20 11
x
y = 36.651 + 2.5528 y = 27.549x - 0.1039
R 2 = 0.5217 10 R 2 = 0.7785
18

9
16
YAN L* (0-24h)
YAN L* (0-24h)

8
14

12
6

10
5

8
4

6 3
0.10 0.20 0.30 0.40 0.10 0.20 0.30 0.40

PPO activity (A 450) PPO activity (A 450)

Figure 2 Relationship between PPO activities and YAN ∆L* (0-24h) in a set of lines from
the DM5637*B8 x H45 DH population in (a) 2005 and (b) 2006 (n=50).
Vol. 43, No.2, 2010 QTL mapping of PPO activity and YAN color in wheat population 113

(a) Year 2005 (b) Year 2006
2
0
0.10 0.20 0.30 0.40
0.10 0.20 0.30 0.40
y = 29.367x - 12.791 1 y=17.748x - 6.7903
-2 R 2 = 0.5277 R 2 = 0.6697
0

-4

YAN b* (0-24h)
-1

YAN b* (0-24h)
-6 -2

-3
-8
-4

-10
-5

-12 -6

PPO activity (A 450) PPO activity (A 450)

Figure 3. Relationship between PPO activity and YAN ∆b* (0-24h) in a set of lines
derived from the DM5637*B8 x H45 DH population in (a) 2005 and (b) 2006 (n=50).

DArT Marker Screening Three QTLs were detected in 2006, together
The DNA of 92 lines and the two parents of the accounting for 70% of variation in PPO activity
DM5637*B8 x H45 population were analysed by (Table 1). A highly significant (P<0.001) QTL on
DArT. Three hundred and eighty four polymorphic chromosome 2AL was also associated with PPO
DArT markers were obtained with an overall call activity in 2006. Marker locus wPt-7024, which
rate of 94.28%. The overall marker P values were was located in the 2A QTL (QPPO.dmh45-2A) had
extremely reliable and ranged between 73.87 and a highly significant association with PPO activity
98.84 with a mean of 89.63. Only 17 markers had P (LRS = 59.9; P>0.001) and accounted for 50% of
values below 77. Of the 384 polymorphic markers the observed variation. The QTL on 7BL
that were identified 317 markers were assembled (QPPO.dmh45-7B) explained 10% of the
into 30 linkage groups using Cartablanche software, phenotypic variation (LRS value of 9.5) and
version 1.5.0(111), Keygene Products B.V. and corresponded with a QTL identified in 2005. In the
QTXb20. The remaining markers were unassigned. 2006 growing season, another QTL (QPPO.dmh45-
7A) was identified on chromosome 7A. It was not
QTL Analyses detected in 2005. Marker locus wPt-0004 within
Polyphenol oxidase activity this QTL was closely associated with PPO activity.
Interval mapping analysis identified three This marker had an LRS value of 8.9 (P>0.01) and
genomic regions that controlled PPO activity in the explained 10% of the variation in PPO activity in
2005 trial (Table 1). A highly significant (P<0.001) 2006. The 4A QTL (QPPO.dmh45-7A) that was
(QTL (QPPO.dmh45-2AL)) was located on chromo- identified in 2005 was not detected in 2006.
some 2AL (LRS=37.2, Figure 4) and explained The three QTLs that were identified in 2006
35% of phenotypic variation. The marker loci wPt- (QPPO.dmh45-2AL; QPPO.dmh45-7A and QPPO.-
5865 and wPt-7024 in the 2AL QTL (QPPO.dmh45- dmh45-7B) were also detected when analysing
2AL) were both significantly associated with PPO pooled data across both seasons (Table 1). The
activity in this season. In 2005, significant QTLs three QTLs together accounted for 71% of the
for PPO activity were also identified on chromo- variation in PPO activity. The association of PPO
somes 4A (QPPO.dmh45-4A) and 7B (QPPO.dmh activity with markers wPt-5865 and wPt-7024 in
45-7B). The three QTLs that were identified the 2AL QTL was highly significant (P<0.001;
collectively explained 53% of the observed LRS of 63.7) and contributed 52% of the observed
phenotypic variation in this season (Table 1). phenotypic variation. A suggestive QTL that was
114 A. Sadeque and M. A. Turner Thai Journal of Agricultural Science

Table 1 Results of interval mapping for PPO activities (A450) in the DM5637*B8 x H45 DH population.

Chromosome Parent
Site-year QTLs Closest marker LRS value1/ PVE (%)2/
location (+ve PPO)
Narrabri-2005 2AL QPPO.dmh45-2AL wPt-7024 37.2** 35 H45
4A QPPO.dmh45-4A wPt-7919 8.9* 10 H45
7B QPPO.dmh45-7B wPt-9925 7.1* 8 H45
Narrabri-2006 2AL QPPO.dmh45-2AL wPt-7024 59.9** 50 H45
4A QPPO.dmh45-4A wPt-0004 8.9* 10 H45
7B QPPO.dmh45-7B wPt-9925 9.5* 10 H45
Pooled 2AL QPPO.dmh45-2AL wPt-7024 63.7** 52 H45
4A QPPO.dmh45-4A wPt-7919 7.4* 8 H45
7B QPPO.dmh45-7B wPt-9925 10.2* 11 H45
1/
Likelihood statistic ratio; * and ** indicate significant and highly significant level at P<0.01 and P<0.001, respectively.
Non asterisk LRS values indicate the suggestive level (P>0.05).
2/
Phenotypic variation explained.

2AL QPPO.dmh45-2AL Q∆L*.dmh45-2AL Q∆b*.dmh45-2AL

63.7 47.7 53.8

Figure 4 Likelihood Ratio Statistic (LRS) and additive effect plots of the QILs on chromosome 2AL for PPO activity,
stability of YAN brightness and yellowness, respectively. LRS scores and additive effect curves (solid and broken
respectively) are shown on the partial maps of chromosome. Vertical lines (green) from left to right indicate suggestive,
significant and highly significant levels of LRS values, respectively.

identified on chromosome 7A (QPPO.dmh45-7A) 2005 (Table 2). The major QTL was located on
(LRS=7.4) explained 8% of phenotypic variation. chromosome 2AL (LRS=48.5) and explained 64%
Analysis of pooled data revealed that the 7B QTL of phenotypic variation. The QTL (Q∆L*.dmh45-
was significantly associated (P>0.01) with PPO 1AS) on chromosome 1AS explained 27% of
activity (LRS=10.2) and explained 11% of phenotypic variation (LRS=13.1). QTL on
phenotype variation. In these QTLs the closest chromosome 1B (Q∆L*.dmh45-1B) and 7AL
identified markers were consistently identified (Q∆L*.dmh45-7A) were also detected. These QTL
across seasons. Interval mapping analysis revealed explained 14% (LRS=7) and 20% (LRS=10.7), of
that H45 contributed alleles of the three QTLs that phenotypic variation, respectively.
conferred elevated PPO activity. In 2006, Q∆L*.dmh45-2AL was the only QTL
detected (LRS value of 30.2) and it explained 47%
Yellow Alkaline Noodle Color Components of variation. Analysis of pooled data detected all of
Interval mapping analysis identified four the four QTLs that were identified in 2005 (Table
genomic regions that controlled ∆L* for YAN in 2). Each was of a similar significance level to that
Vol. 43, No.2, 2010 QTL mapping of PPO activity and YAN color in wheat population 115

Table 2 Results of interval mapping of yellow alkaline noodle (YAN) color stability CIE ∆L* and ∆b* at 0-24 hours in
the DM5637*B8 x H45 DH population.

Chromosome Contribution
Site-year QTLs Closest marker LRS value1/ PVE (%)2/
location parent
Narrabri-2005
YAN ∆L* 1AS Q∆L*.dmh45-1AS wPt-6455 13.1** 27 H45
1B Q∆L*.dmh45-1B wPt-0705 7* 14 DM5637*B8
2AL Q∆L*.dmh45-2AL wPt-7024 48.5** 64 DM5637*B8
7AL Q∆L*.dmh45-7AL wPt-0004 10.7* 20 H45
7BL Q∆a*.dmh45-7BL wPt-4140 6.8* 13 H45
YAN ∆b* 1AS Q∆b*.dmh45-1AS wPt-6455 7.6* 17 H45
2AL Q∆b*.dmh45-2AL wPt-7024 47.1** 63 DM5637*B8
7AL Q∆b*.dmh45-7AL wPt-0004 12.1* 22 H45
Narrabri-2006
YAN ∆L* 2AL Q∆L*.dmh45-2AL wPt-7024 30.2** 47 DM5637*B8
YAN ∆b* 2AL Q∆b*.dmh45-2AL wPt-7024 39.2** 57 DM5637*B8
7AL Q∆b*.dmh45-7AL wPt-0004 8.9* 17 H45
Pooled
YAN ∆L* 1AS Q∆L*.dmh45-1AS wPt-6455 12.5** 26 H45
1B Q∆L*.dmh45-1B wPt-0705 7.1* 14 DM5637*B8
2AL Q∆L*.dmh45-2AL wPt-7024 47.7** 64 DM5637*B8
7AL Q∆L.dmh45-7AL wPt-3226 8.8* 17 H45
YAN ∆b* 1AS Q∆b*.dmh45-1AS wPt-6455 7.7* 17 H45
2AL Q∆b*.dmh45-2AL wPt-7024 53.8** 68 DM5637*B8
7AL Q∆b*.dmh45-7AL wPt-0004 12.3* 23 H45
1/
Likelihood statistic ratio; * and ** indicate significant and highly significant level at P<0.01 and P<0.001, respectively.
Non asterisk LRS values indicate the suggestive level (P>0.05).
2/
Phenotypic variation explained.

for year 2005. Interval mapping analysis revealed a This QTL had a positive additive effect which was
positive additive affect of the Q∆L*.dmh45-1B and significantly associated with brightness stability of
the major QTL (Q∆L*.dmh45-2A) on chromosome yellow alkaline noodles that was contributed by the
2AL which indicated that parent DM5637*B8 parent DM5637*B8. Regression analysis identified
contributed a positive allele at each of these QTLs. that marker wPt-7024 was significantly (P<0.001)
The additive effect for the other QTLs associated with ∆b*. The other significant QTL on
(Q∆L*.dmh45-1AS and Q∆L*.dmh45-7AL) were chromosome arm 7AL was detected across the
negative in both growing seasons, indicating that growing seasons and accounted for 22% and 17%
parent H45 contributed the positive allele for the of phenotypic variation (LRS=12.1 and 8.9), in
QTLs. 2005 and 2006, respectively. This QTL explained
QTL for YAN trait ∆b* were detected on 23% of variation across seasons. The marker wPt-
chromosomes 1AS (Q∆b*.dmh45-1AS), 2AL 0004 was closely linked to locus controlling this
(Q∆b*.dmh45-2AL) and 7AL (Q∆b*.dmh45-7AL) QTL. The negative additive effect recorded for
were detected (Table 2). Q∆b*.dmh45-1AS was QTLs located on chromosomes 1AS and 7AL
detected only in 2005 and explained 17% of indicate that the high PPO parent (H45) contributed
observed variation in that season (LRS=7.6). The the positive allele for these QTLs. In pooled
QTL on 2AL (Figure 4) and 7AL were detected in environments, the three QTLs (Q∆b*.dmh45-1AS,
both years. The 2AL was highly significant Q∆b*.dmh45-2AL and Q∆b*.dmh45-7AL) were
(P<0.001) in both years, accounting for 63% and significant (P<0.01) explaining 17%, 68% and 23%
57% of variation in ∆b* (with LRS values of 47.1 of phenotypic variation, respectively.
and 39.2 in 2005 and 2006, respectively, Figure 4).
116 A. Sadeque and M. A. Turner Thai Journal of Agricultural Science

Discussion using DArT markers. A large amount of phenotypic
variation was explained by the QTLs for changes in
Both ∆L* and ∆b* were significantly (P<0.01) brightness, Q∆L*.dmh45-2AL, and yellowness,
correlated with PPO activity across growing Q∆b*.dmh45-2AL, (64% and 68% respectively) on
seasons. The findings of this study are in agreement chromosome 2A. Low PPO wheat line
with the reports of the role of PPO on time DM5637*B8 exhibited color stability in both years.
dependent darkening (darkening due to aging) on Not many studies concerned with mapping of color
yellow alkaline noodles (Kruger et al., 1994; Morris stability of YAN in Australian wheat have been
1995; Fuerst et al., 2006). conducted and little information is available. Mares
The results of the current study reconfirm that and Campbell (2001) found a QTL on chromosome
PPO activity in wheat is mainly controlled by a 2D that was clearly associated with these traits and
locus on the long arm of chromosome 2A with another on 2A which had some association. These
strong genetic association with DArT marker wPt- findings are partly in agreement with findings of
7024. Other studies have identified that PPO the current study. The marker locus wPt-7024 was
activity is strongly associated with the SSR marker identified as an important DArT marker for all of
loci Xgwm312, Xgwm294 and WMC170 which are the color stability traits of YAN, and PPO activity.
located on chromosome 2A (Sun et al., 2005; Correlations between PPO activity and change
Raman et al., 2005; Watanabe et al., 2006). The in brightness were observed in this population.
chromosome 2A location is consistent with QTL mapping detected that DArT marker locus
previous findings (Sun et al., 2005; Raman et al., wPt-7024 on chromosome 2AL had the closest
2007) however it is not certain if the identified genetic association with PPO activity and color
QTLs are in precisely the same location as stability of both ∆L* and ∆b* (52% and 68% PVE,
identified in previous studies. The association of respectively). QTL mapping results identified
DArT marker wPt-9925 on chromosome 7BL with common locations of QTLs for both characters.
PPO activity in wheat has not yet been reported in Also the parental contribution was consistent in
the literature. This QTL (QPPO.dmh45-7BL) each case. The DArT marker locus wPt-7024 had
explained 8-11% phenotypic variation across the highest contribution to YAN color components
environments in consecutive years. QTLs on ∆L* and ∆b*. Therefore, it can be concluded that
chromosomes 4A and 7A were detected in one PPO activity is the major factor that influences
season (Narrabri 2005 and 2006, respectively). The YAN color stability. DArT marker wPt-7024 was
QTL on 7A, was also detected when data was associated with both YAN brightness and
pooled across environments. A QTL on yellowness along with PPO activity. In addition,
chromosome 7A was also detected in other studies two potential QTLs for YAN ∆L* and ∆b* have
involving hexaploid wheat that were grown at the been identified on chromosomes 1AS and 7AL.
same site (Sadeque 2008). Therefore, it is possible The QTL on 7AL was detected across
that the minor QTL that is located on environments. A QTL on chromosome 7A was also
chromosmome 7A is sensitive to environmental identified for PPO activity strengthening the
influence. The QTL QPPO.dmh45-4A explained hypothesis that it is an important determinant of the
10% of variation in PPO activity in one season change in YAN brightness. It is interesting that a
(Narrabri 2005). There are no published reports of locus on 7B for PPO activity was identified in both
the presence of QTL on chromosome 4A, thus the seasons but a corresponding QTL for change in
existence of QTL on chromosome 4A needs to be brightness was not detected in this analysis.
confirmed. The analyses of allelic effects of the The identification and mapping of PPO genes in
QTLs revealed that selection using markers within wheat is an important prerequisite for developing
QPPO.dmh45-2AL may be an efficient means to low PPO wheat cultivars that exhibit high levels of
select low PPO activity genotypes in this DH color stability in wheat end products, especially for
population. yellow alkaline and white salted noodles. This
In the present study, QTL mapping of stability study confirms that PPO activity in wheat is under
of YAN color components were also performed polygenic control. The DArT marker wPt-7024 that
Vol. 43, No.2, 2010 QTL mapping of PPO activity and YAN color in wheat population 117

is located within a QTL on chromosome 2AL and Fuerst, E.P., J.V. Anderson and C.F. Morris. 2006.
and has a high level of significance with PPO Delineating the role of polyphenol oxidase in the
darkening of alkaline wheat noodles. J. Agric. Food
activity (P<0.001). In this study a new QTL Chem. 54: 2378-2384.
location on chromosome 7B was identified. A He, X.Y., Z.H. He, L.P. Zhang, D.J. Sun, C.F. Morris,
possible QTL location on chromosome 7A was also E.P. Fuerst and X.C. Xia. 2007. Allelic variation of
revealed in this study. The identification of these polyphenol oxidase (PPO) genes chromosome 2A
and 2D and development of functional for the PPO
QTLs and markers associated with PPO activity in
genes in common wheat. Theor. Appl. Genet. 115:
this study will enable application of marker assisted 47-58.
selection in plant breeding programs of Eastern Huttner, E., P. Wenzyl, J. Carling, L. Xia, S. Yang, D.
Australia which is an efficient approach to pyramid Jaccoud, V. Caig, M. Evers, G. Uszynsi, N. Howes,
genes for low PPO activity. The results of the P. Sharp, P. Vaughan, B. Rathmell and A. Kilian.
2006. Triticarte: whole-genome profiling service for
current study confirmed earlier findings about the wheat and barley using Diversity Arrays Technology
involvement of some genomic regions in (DArT). Proceedings 13th Australasian Plant Breeding
controlling the traits investigated. In addition, new Conference. 18-21 April, 2006. Christchurch, New
genomic regions and associated DArT markers Zealand.
Jaccoud, D., K. Peng, D. Feinstein and A. Kilian. 2001.
were identified that were linked to loci that control
Diversity Arrays: a solid state technology for
wheat PPO and YAN color stability components. sequence information independent genotyping. Nucl.
Acids Res. 29: e25.
Acknowledgments Jimenez, M. and J. Dubcovsky. 1999. Chromosome
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CRC Ltd. genetic testing, pp. 27-44. In G.B. Fincher and C.
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Genetic mapping of the gene affecting polyphenol
Manuscript received 3 March 2010, accepted 30 September 2010

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