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Working in Partnership to Attain Priority Crop Genetic Resource Genomics and Genetic Improvement Research Goals PETER BRETTING U S DA A G R I C

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Working in Partnership to Attain Priority Crop Genetic Resource Genomics and Genetic Improvement Research Goals PETER BRETTING U S DA A G R I C Powered By Docstoc
					   Working in Partnership to Attain Priority
Crop Genetic Resource, Genomics, and Genetic
        Improvement Research Goals




                         PETER BRETTING
     U S DA - A G R I C U LT U R A L R E S E A R C H S E R V I C E
             O F F I C E O F N AT I O N A L P R O G R A M S
            PETER .BRET TING@ARS.USDA .GOV
  H T T P : / / W W W. A R S . U S D A . G O V/ R E S E A R C H / P R O
      GRAMS/PROGRAMS.HTM?NP_CODE=301
                    Outline for Presentation


 American Seed Research Summit
 USDA/ARS National Program in Plant Genetic Resources,
 Genomics, and Genetic Improvement
    Plant Genetic Resources and Information Management
    Plant Genomics, Genetic Analyses and Genome Databases
        Maize and soybeans: genotyping and nested association mapping
    Genetic Improvement of Crops
        Maize: GEM Project
 Conclusion
              American Seed Research Summit
      Research, Education, and Policy Goals and Strategies

 Strengthen public and private partnerships to accomplish
    national seed research priorities
   Coordinate and engage industry stakeholders to support stable
    funding for seed and breeding education, research, and
    development
   Attract and develop a pool of diverse, high-quality plant
    researchers
   Ensure that the regulatory system governing the development
    and implementation of new technology is efficient, effective,
    and science-based.
   Develop an education and advocacy program to communicate
    the value of seed and crop research to the public.
   USDA/ARS National Program in Plant Genetic Resources,
   Genomics, and Genetic Improvement (Crop Genes R’ Us!)



 Largest NP, with 125+         Goal: deliver crop genetic,
  projects, and about $128       genomic and bioinformatic
  million gross annual           tools, information, genetic
  budget.                        resources, and improved crop
                                 varieties to enhance U. S.
 Conducted by about 300         agricultural productivity and
  scientists at more than 50     security.
  ARS locations.
 Extensive public and
  private sector
  partnerships.
   NP 301 Action Plan Research Components


 Plant and Microbial Genetic Resource
 Management

 Crop Informatics, Genomics, and Genetic
 Analyses

 Genetic Improvement of Crops
   The USDA/ARS National Plant Germplasm System
                     (NPGS)



 One of the largest national    Large holdings of crops
  genebank systems.               without major collections at
                                  international agricultural
 More than 510,000 samples       research centers, e.g.,
  of more than 13,400 plant       cotton, soybean, various
  species.                        horticultural and “specialty”
 Large collections of the        crops.
  major staple crops             Germplasm Resources
  important to U. S. and          Information Network
  world agriculture.              (GRIN): an international
                                  standard.
USDA National Plant Germplasm System (NPGS)
Plant Genetic Resource Management in Genebanks


  Acquisition          Characterization
  Maintenance          Evaluation
  Regeneration         Enhancement
  Documentation and
   Data Management
  Distribution
Avoidance of Cross Pollination and Seed Mixing


 Extensive quality assessment when the seed enters
  the NPGS
 Procedures and protocols for regeneration
    Physical isolation
    Cages
    Hand pollination
 Care in storage and distribution to prevent mixing
              Maize in the NPGS

 Isolation from sources of out
  crossing
 Regenerate ~400 accessions/year
 Sometimes plantings yield as few
 as 5,000 seeds for some accessions
                        GRIN-Global

 GRIN = Germplasm Resources Information Network.
  http://www.ars-grin.gov/ The genebank information
  management system for the NPGS, and for Canada’s genebank
  system (GRIN-Canada).
 The Global Crop Diversity Trust asked ARS and Bioversity
  International (an International Agricultural Research Center) to
  enhance and expand GRIN to address global germplasm
  information management needs.
 The Trust awarded ARS a 3-year, $1.4 million grant to develop
  GRIN-Global; ARS is devoting $900K in-kind support to the
  project. ARS effort is located in Beltsville, MD and Ames, IA.
                     GRIN-Global

 Based on GRIN, but can be implemented in both a
  system-wide and “stand-alone” local management
  mode
 Supports multiple users via a “user-friendly” interface
 Maintains linkages with other databases and
  interoperates with existing systems
 Advanced querying, custom and third-party
  applications
  Three Tier Architecture

                                 Presentation
 Presentation                        Tier
     Tier                         (Windows
(Web Browser)                   Desktop Client)




                Business Tier
                (Web Service)




                 Data Tier
                 (MySQL,
                Oracle, SQL
                  Server)
                      GRIN-Global


 On-line ordering/request capability
 Database-flexible, free of recurrent licensing costs, with
  interface and database schema source code open and
  available without restriction to further development
 Designed to serve as the global standard plant genebank
  information management system
Future Prospects
 Trends in demand for NPGS germplasm and information
                   vs. NPGS budget

        NPGS Web Page Access                                                                  Germplasm                                                                       NPGS Budget
2,500,000                                                                                     Distributions                                                      45,000,000
                                                                                  200000                                                                         40,000,000
2,000,000                                                                         180000
                                                                                                                                                                 35,000,000
                                                                                  160000
                                                                                                                                                                 30,000,000
1,500,000                                                                         140000
                                                                                  120000                                                                         25,000,000
                                                                                  100000                                                                         20,000,000
1,000,000
                                                                                   80000
                                                                                                                                                                 15,000,000
                                                                                   60000
 500,000                                                                                                                                                         10,000,000
                                                                                   40000
                                                                                   20000                                                                          5,000,000

       0                                                                               0                                                                                 0
            1999
                   2000
                          2001
                                 2002
                                        2003
                                               2004
                                                      2005
                                                             2006
                                                                    2007
                                                                           2008




                                                                                           1999
                                                                                                  2000
                                                                                                         2001
                                                                                                                2002
                                                                                                                       2003
                                                                                                                              2004
                                                                                                                                     2005
                                                                                                                                            2006
                                                                                                                                                   2007
                                                                                                                                                          2008




                                                                                                                                                                              1999
                                                                                                                                                                              2000
                                                                                                                                                                              2001
                                                                                                                                                                              2002
                                                                                                                                                                              2003
                                                                                                                                                                              2004
                                                                                                                                                                              2005
                                                                                                                                                                              2006
                                                                                                                                                                              2007
                                                                                                                                                                              2008
Result?: mismatch between expanding demand for PGR and static NPGS
                       capacity to manage it.
   NP 301 Action Plan Research Components


 Plant and Microbial Genetic Resource
 Management

 Crop Informatics, Genomics, and Genetic
 Analyses

 Genetic Improvement of Crops
    Genomic Information and Research Tools


 Genetic markers: polymorphic and heritable—simple
  sequence repeats (SSRs) and single nucleotide
  polymorphisms (SNPs)
 Expressed sequence tags (ESTs)
 Genetic maps
 Quantitative trait loci (QTLs)
 Physical maps
 Complete and partial genome sequences
Application of Genomic Information to Genetic Analyses:
The Nested Association Mapping (NAM) Genetic Map

[E. Buckler, J. Holland, M. McMullen (USDA/ARS), and many university and
private-sector collaborators]

NAM is the most powerful tool for dissection of the genetic bases of
quantitative traits for any species – period.
Linkage Mapping                           Association Mapping

Recent recombination                            Historic recombination
High power                                                  Low power
Low resolution                                          High resolution
Analysis of 2 alleles                          Analysis of many alleles
Moderate marker density                            High marker density
Genome scan                                     Candidate gene testing


               Nested Association Mapping
                 Recent and ancient recombination
                            High Power
                          High resolution
                      Analysis of many alleles
                 Moderate genetic marker density
                   High projected marker density
                                           NAM
                                                                     F1s



                                                         SSD
                                                                                           25 DL




                                           
                                           
                                           
                                                 2
                                                     1




                                     200
                                                                                    B97




                                                         
                                                             
                                                                 
                                                                                 CML103
                                                                                 CML228
                                                                                 CML247
                                                                                 CML277




                                                             
                                                                                 CML322




                                                             
                                                                                 CML333




                                                             
                                                                                 CML52
                                                                                 CML69
                                                                                  Hp301
                                                                           ×




                                                                                   Il14H
                                                                           B73




                                                                                   Ki11
                                                                                    Ki3
                                                         
                                                             
                                                                 




                                                                                   Ky21
                                                                                 M162W
                                                                                  M37W
                                                                                 Mo18W
                                                             




                                                                                  MS71
                                                             




                                                                                  NC350
                                                             
                                                                                                   Nested Association Analysis




                                                                                  NC358
                                                                                   Oh43
                                                                                  Oh7B
                                                                                    P39
                                                                                  Tx303
                                                                                   Tzi8
                                                         
                                                             
                                                                 




Yu et al. (2008) Genetics 178: 539
    NAM Genotyping and Genetic Map

•Genotyping with more than 1500 single nucleotide
polymorphism (SNP) genetic markers

•Map consists of 1106 loci (38 composite loci) and ~1400
cM genetic distance, therefore an average marker density of
1.3 cM/marker.

•It is a composite or consensus map and distances and
potentially order can not be assumed to translate directly
to individual family maps.
                  Maize Phenomics:
Massively Parallel Phenotyping of The Nested Association
                   Mapping Population




       THE MAIZE DIVERSITY PROJECT
          Experimental Evaluation of NAM
     NAM lines evaluated in 2006 – 2007 at each of the following sites:




                                                        Aurora, NY

                                     Champaign, IL
                     Columbia, MO
                                                                Clayton, NC



                                                    Homestead, FL

                                                                          Ponce, PR
11 Total Environments
           +


                        Tassel main spike length
Tassel branch angle
                      Tassel branch no.
                      Partitioning Genetic Variance

    B73                   B73      B73          B73          B73                 B73
     X                     X        X            X            X                   X
   NC358                  P39     M37W         IL14H        Tx303                B97
 B73
                Line-to-line variation within a family – due to
                 B73        B73       B73        B73       B73
  X             differences X
                  X          between alleles from B73 and X
                                       X          X
Mo18W           Ki11       NC350                Oh7B
                alleles from DiverseKy21 founder of that Ki3
                                      Line
     B73        family.B73       B73                  B73                        B73
                                           B737
      X                X           X                         X                    X
                                             xHp301
    CML103            Oh43        MS71                     M162W                 Tzi8

  B73        B73             B73        B73             B73          B73                 B73
   X          X               X          X               X            x                   X
 CML52      CML322          CML228     CML277          CML247       CML333              CML69



 RIL1    RIL2    …          Among-Family Genetic Variance: σ2F
                      RIL199 RIL200                       RIL1 RIL2  …RIL         199    RIL200
                            Family-to-family variation – due to
                            Within-Family Genetic Variances
                            differences among different founders.
                σ2G(F)1                                             σ2   G(F)2
                             The Maize Genetics and
                          Genomics Database (Maize GDB)
                               (C. Lawrence et al.)
                     USDA/ARS, AMES, IA www.maizegdb.org/

 Phenotypes
 Integrating structural and genetic
  maps with maize genomic sequence
 Genome annotation
 Research community support
Soybean Nested Association Mapping (NAM) Project
       (P. Cregan, D. Hyten, ARS-Beltsville)

    Collaborators:
    USDA/ARS
   Iowa St. Univ.
 Univ. of Nebraska
   Univ. of Illinois
 Univ. of Tennessee
 Univ. of Maryland

   Agricultural
   Research
   Service
                  Funding Support
                      Soybean NAM
 Design focused on yield
   Populations will be restricted to Maturity Group III

 Hub parent is IA3023
 Lines crossed with IA3023 included both elite and exotic
  germplasm
     121 total lines selected by research community
      –   Crosses made the summer of 2008

 Parents were genotyped with the Universal Soy Linkage
  Panel 1.0
  What’s next with the Soybean NAM?
 During population development, the number of families
 will be reduced to 40 based on field observations during
 the inbreeding process.
    Each population will consist of 250 F5 RILs
     –   Total size: 10,000 lines

 Phenotyping will be done with a ‘connected’ incomplete
 block experimental design (Tested in 30 environments)
    Best Linear Unbiased Predictions (BLUPs) of grain yield, i.e., breeding
     values for every RIL
  The Universal Soy Linkage Panel 1.0 (USLP 1.0)


With United Soybean Board and collaborator funding 180 sets of
 the USLP 1.0 (17,280 genotypes) were acquired for gene/QTL
                          Discovery

ARS Collaborators                 State University
    Ames, IA                       Collaborators
  Beltsville, MD
                          N. Carolina St.      Univ. of Georgia
  Columbia, MO
                                Univ.           Univ. of Illinois
   Raleigh, NC          N. Dakota St. Univ.   Univ. of Minnesota
  Stoneville, MS         Ohio State Univ.      Univ. of Missouri
    Urbana, IL           So. Illinois Univ.   Univ. of Nebraska
  Wooster, OH           S. Dakota St. Univ.     Virginia Tech
                        Univ. of Arkansas
     Universal Soy Linkage Panel (USLP 1.0)
1,536 SNPs selected from 3,110 SNPs mapped on the
 Soybean Consensus Map
     SNPs have diverse allele frequencies
        Average polymorphism in bi-parental crosses
          Elite cultivars= 458

          PI landraces = 544

          Elite crossed with PI landrace = 590

    Spread throughout the genome
    Assayed using the Illumina GoldenGate assay
        192 DNA samples run in three days
        Large orders reduce cost from $11,000 per 96 DNA samples to $5,500
         per 96 DNA samples
       The Universal Soy Linkage Panel 1.0 (USLP 1.0)
                  for Gene/QTL Discovery

        - Traits Under Study in Collaborative Projects -

Enhanced Seed             Resistance to             Resistance to
 Composition              Biotic Stress             Abiotic Stress

Reduced linolenic acid    Soybean Cyst Nematode
                                                        Drought
         oil                   Soybean Rust
                           Phytophthora Root Rot    Iron Deficiency
Elevated oleic acid oil
                           Foliar Feeding Insects      Chlorosis
   Lower saturates             Soybean Aphid
    Higher protein




  Seed Yield – Assessment and enhancement of genetic diversity
Source of Genetic Improvement for Soybean
 USDA germplasm collection is the source for new genetic
  variation for soybean improvement for every important trait
          USDA Soybean Collection (21,000 accessions)




                                                 18,090 accessions
     1,116 wild                                   collected mostly
     soybeans                                         from Asia
                                                    (Landraces)



   17 Introduced                                      513 Public
     landraces                                         cultivars
    (86% of the           Hyten et al. 2006, PNAS 103: 16666-16671
                                                    released after
  Germplasm base)                                        1946
                            Soy HapMap
• The United Soybean Board recently agreed to fund a $2.9M
    project to characterize the entire USDA Soybean Germplasm
    Collection of 21,000+ wild and cultivated soybeans with
    50,000 SNP DNA markers.
•   The Illumina’s Beadstation will allow genotyping of this
    collection with 50,000 SNPs within three years
    • New gene discovery through association analysis
    • Enables breeders to select germplasm with greatest potential for
        agronomic improvement
    •   Decipher the signatures of selection (allele frequency changes)
        associated with soybean yield improvement over 75 years of soybean
        breeding to help understand yield

Soybean Genomics and
  Improvement Lab
USDA, ARS, BARC-West
 Beltsville, Maryland      60,800 SNP Infinium Chip
 Research by R. Shoemaker (ARS-Ames), C. Vance (ARS-St. Paul) and
  collaborators at N. Dakota State Univ. and Iowa State Univ.
 Iron is a limiting growth factor on 30% of cropland
      Iron is also a major nutritional deficiency in much of the world

 Using global gene expression profiling to identify genes involved in iron
  metabolism
 Currently evaluating how to control the genetic expression to enhance iron
  balance in commercial plant varieties.
 Developing molecular markers to speed the public release of commercial
  varieties with improved iron efficiency.
Mapping & Cloning Genes Responsible for Soybean
                    Protein


 Research by R. Shoemaker (ARS-Ames), C. Vance
  (ARS-St. Paul), collaborators at the Univ. of Illinois
  and Univ. of Nebraska
 The locus of a major trait that controls                                 Soybean Seed
  seed protein content was mapped                          to              Composition
  Chromosome 20
 High-throughput transcript                          15% Soluble                          18% Oil
                                                     Carbohydrates
  sequencing and mapping identified               several
                                                    (sucrose, stachyose,
                                                      raffinose, others)
  candidate genes
                                                                                          14% Moisture,
 Currently identifying the specific               genes                                    ash, other
  and evaluating modifications
  to change the protein content                         38% Protein
                                                                                          15% Insoluble
                                                                                          Carbohydrates
SNP Haplotypes and Linkage Disequilibrium
           (R. Shoemaker, ARS-Ames)


                      Haplotype and Allele States for varieties
   NP 301 Action Plan Research Components


 Plant and Microbial Genetic Resource
 Management

 Crop Informatics, Genomics, and Genetic
 Analyses

 Genetic Improvement of Crops
Germplasm Enhancement of Maize
                                 Germplasm Enhancement of Maize (GEM) Project


                                    A collaborative effort of public and private sector
                                    researchers to broaden and enhance the maize germplasm
                                    base. More than 60 collaborators.
                                    Two permanent breeding sites:
                                      Ames, IA for development of 25% tropical and temperate exotic
                                      Raleigh, NC for development of 50% tropical
                                    GEM is administered by the USDA-ARS Plant Introduction
                                    Research Unit (PIRU) located in Ames, IA; and the Plant
                                    Science Research Unit (PSRU) in Raleigh, NC
                                    Technical Steering Group (TSG) provides guidelines for
                                    research, germplasm, and methods.
Germplasm Enhancement of Maize
                                 GEM Project Cooperators


                                 Category                          Number
                                 Private US companies                29
                                 Public US universities              13
                                 USDA-ARS research units             8
                                 Non-government organization         1
                                 Private international companies     11
                                 Public international institutes     4
                                 Total                               66
Germplasm Enhancement of Maize
                                         GEM Objectives


                                 Manage and coordinate a multi-site cooperative
                                 program for germplasm evaluation, development,
                                 and information sharing
                                 Evaluate diverse maize germplasm for adaptation,
                                 yield, stress resistance, and key value-added traits
                                 (VATs)
                                 Develop and release enhanced germplasm with
                                 key traits
                                 Develop innovative means of managing and
                                 transferring information to the maize community
Germplasm Enhancement of Maize                            GEM Germplasm Releases
                                  Year       # Lines Released            Institution                   Germplasm Attributes
                                 2001                  1*                USDA-ARS Ames                 Resistance to 1st brood ECB (non-DIMBOA)
                                 2002                  2*                Univ. of Delaware             Yield, resistance to anthracnose and GLS
                                 2003                 29**               NC State Univ.                Yield, earlier flowering, GLS, Fusarium Resistance
                                 2003                  1*                Ohio State Univ.              Yield, Fusarium resistance
                                 2003                  1                 Univ. of Delaware             VAT
                                 2003                  42                USDA-ARS Ames                 Temperate adaptation, GLS, VAT
                                 2003                  9*                NC State Univ.                Yield, VAT, GLS
                                 2004                  14                USDA-ARS Ames                 Temperate adaptation, yield, VAT
                                 2004                  2                 Texas A&M Univ.               Stress tolerance, yield, CEW, grain mold resistance
                                 2004                  1                 Univ. of Wisconsin            Superior nutritional quality/yield
                                 2004                  9                 NC State Univ.                Yield, earlier flowering, VAT
                                 2005                  9                 USDA-ARS Ames                 Temperate adaptation, yield, VAT
                                 2005                  1                 Univ. of Delaware             High protein
                                 2005                  19                NC State Univ.                Yield, earlier flowering, VAT
                                 2006                  13                USDA-ARS Ames                 Yield, VAT
                                 2006                  3                 NC State Univ.                Yield, earlier flowering
                                 2007                  10                USDA-ARS Ames                 Protein, oil, high starch for ethanol
                                 2007                  10                NC State Univ.                50% exotics; disease resistance
                                 2007                  1*                Truman State Univ.            Amylose maize VII line (GEMS-0067)
                                 2008                  13                USDA-ARS Ames                 Temperate adaptation, yield, VAT, waxy lines
                                 2009                  7                 USDA-ARS Ames                 Protein, oil
                                 2009                  5                 USDA-ARS Raleigh              50% exotic; protein, oil
                                 Total                202
                                 * Crop Science registered and ** 20 of these 29 lines were Crop Science registered.
Germplasm Enhancement of Maize
                                  Number of Releases by Traits

                                 Germplasm Traits                            Number
                                 Protein above 13%                             33
                                 Amino Acid index (Met. Lys. Trp.)             17
                                 Oil above 4.5%                                13
                                 Starch compositional properties               24
                                 High Amylose (70%)                            1
                                 Waxy                                          2
                                 Fumonisin (reduced level)                     5
                                 Aflatoxin (reduced level)                     6
                                 Leaf disease resistance (GLS, NLB, & SLB)    104
                                 Other (yield, Y/M, etc.)                      12
Germplasm Enhancement of Maize
                                        New GEM Initiatives


                                 Allelic diversity
                                   New un-sampled races being tapped
                                   Goal to assess ~300 races for adaptation in US
                                 New elite exotic sources being acquired
                                   More than 40 germplasm sources acquired from
                                   Thailand, Peru, Nigeria, Argentina, Chile, and France
                                   Provide resistance to exotic diseases
                                 Shade house to reduce photoperiod response
                                   Make tropical introgressions in Ames, IA
                                   23 tropical sources (11 races) successful (so far)
                                 Double haploid research
                                   Explore feasibility with exotic germplasm
                                         Fusarium Ear Rot
Germplasm Enhancement of Maize




                                 Susceptible Line                    Resistant Line
                                                                    GEMS-0002 Public Release

                                                    Bill Dolezal, Pioneer Hi-Bred Int’l, Woodland, CA (2005)
Integrated national programmatic approach, with extensive
  academic and private sector collaborations/partnerships
            Exploit untapped genetic diversity in genebanks, and breeding
            populations

                     Plant genomics, gene discovery


                                 Genetic markers & Bioinformatics


                                        Genome Databases


                                                           Genetic
                                                         Enhancement
                                                         and Breeding
              American Seed Research Summit
      Research, Education, and Policy Goals and Strategies

 Strengthen public and private partnerships to accomplish
    national seed research priorities
   Coordinate and engage industry stakeholders to support stable
    funding for seed and breeding education, research, and
    development
   Attract and develop a pool of diverse, high-quality plant
    researchers
   Ensure that the regulatory system governing the development
    and implementation of new technology is efficient, effective,
    and science-based.
   Develop an education and advocacy program to communicate
    the value of seed and crop research to the public.
                    Thanks!


 Thanks to the NCCPB and ASTA for the invitation
  to speak
 Thanks to USDA/ARS scientists for sharing
  information and data
 Thanks to our partners in the seed industry and
  academia for their invaluable collaboration

				
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