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United States Department of Agriculture

Agricultural Research Service

Research Project: DEFINING THE GENETIC DIVERSITY AND STRUCTURE OF THE SOYBEAN GENOME AND APPLICATIONS TO GENE DISCOVERY IN SOYBEAN AND WHEAT GERMPLASM

Location: Soybean Genomics and Improvement

2010 Annual Report


1a.Objectives (from AD-416)
The three objectives of the research are firstly, to define linkage disequilibrium and recombination rates across the soybean genome to facilitate efficient discovery of quantitative trait loci (QTL) through Association Analysis and efficient introgression of exotic germplasm, secondly, to define genome regions in cultivated soybean that are associated with domestication for the discovery of genetic variation lost through the domestication bottleneck that can be used to improve soybean and thirdly, to discover QTL and genes controlling biotic and abiotic stress resistance and quality traits in soybean and wheat, and develop DNA markers that define haplotype variation across these and previously identified regions.


1b.Approach (from AD-416)
Single nucleotide polymorphism (SNP) DNA markers will be discovered using high throughput genome sequence analysis in combination with the newly developed whole genome soybean sequence from the Department of Energy, Joint Genome Institute. A set of 50,000 SNPs, selected from across the genome, will be identified and genetically mapped in cultivated soybean as well as in a newly created cultivated x wild soybean population. The same SNPs will be used to characterize 16,795 soybean landraces as well as a set of 96 elite soybean cultivars and 1,116 wild soybean genotypes. This will allow an assessment of linkage disequilibrium and population structure across the genomes of the landraces, elite cultivars and wild soybeans. Association Analysis will be assessed as a new approach to detect genes/QTL underlying the important trait of seed protein concentration. The high resolution genetic maps in both cultivated x cultivated and cultivated x wild soybean populations combined with QTL analysis of traits related to soybean domestication will facilitate the identification of regions in cultivated soybean which, in comparison to wild soybean, have little or no genetic variation as a result of “selective sweeps” that occurred during soybean domestication. A universal set of 1536 soybean SNPs with high rates of polymorphism and even distribution across the genome will be developed and used to discover QTL underlying a number of disease resistance and quality traits in soybean. In addition, DNA marker development in hexaploid wheat will be continued and these markers and other SSR markers previously developed in our laboratory will be used in QTL analysis for a number of important traits in hexaploid wheat.


3.Progress Report
Progress was made in development of the Soybean Illumina Infinium beadchip which is capable of analyzing 50,000 single nucleotide polymorphism (SNP) DNA markers in the DNA of the soybean genome. The 50,000 SNPs were selected from among approximately 200,000 SNP markers developed using “next generation” Illumina Genome Analyzer DNA sequencing of a set of six cultivated and one wild soybean. The sequences were aligned to each other and to the newly completed whole genome sequence of the soybean for SNP discovery and to determine the position of each putative SNP on the 20 soybean chromosomes. The 50,000 SNPs were then selected from the total of 200,000 SNPs based upon their position on the 20 soybean chromosomes to assure an even distribution of SNP markers across each of the 20 soybean chromosomes. The Soybean Illumina Infinium Beadchip was used to analyze more than 10,000 accessions from the USDA Soybean Germplasm Collection. The analyzed accessions included accessions with high seed protein concentration.

Progress was made in discovery of genes controlling the level of protein in the soybean seed. In collaborative research, 48 soybean germplasm accessions with high seed protein (48% or more) were mated to a high-yielding cultivar with normal seed protein (42% or less) of the same maturity to generate the 48 populations. A total of 240 F2 plants per mating were grown in the field and a leaf sample was collected from each for DNA isolation. The F3 seeds from the 240 plants from each mating were analyzed for seed protein concentration. Using a techniques called “selective genotyping” DNA from the 22 F2 plants that produced the seed with the highest seed protein concentration and the 22 plants that produced the seed with the lowest seed protein concentration from each mating were analyzed along with the two parental lines with the 1536 SNP DNA markers in the Universal Soybean Linkage Panel 1.0. To date significant genetic effects controlling seed protein that have not been previously reported have been detected on five different soybean chromosomes.

Progress was made in the discovery of SNP DNA markers in wheat using next generation Roche 454 and Illumina Genome Analyzer DNA sequencing. Genomic DNA libraries were constructed by restriction enzyme digestion and size selection of DNA fragments in the 400-600 basepair size range of two hard red winter wheats, two hard red spring wheats two soft red winter wheats and one durum wheat. These libraries were further digested and sub-libraries with DNA fragments in the 180-240 basepair size range were isolated. The 400-600 basepair fragments of the library of the cultivar Chinese Spring were sequenced with the Roche 454 DNA sequencer and each of the seven sub-libraries were sequenced with the Illumina Genome Analyzer. The resulting DNA sequences are being aligned and analyzed to identify SNP DNA markers in preparation for the design of a 6,000 SNP Illumina Infinium beadchip.


4.Accomplishments
1. Discovery, evaluation, and release of 33,065 Simple Sequence Repeat (SSR) DNA markers in soybean. DNA markers serve as genetic landmarks and are interspersed among and within the genes throughout the genome of higher organisms including the soybean. If a marker is located near a gene of interest, the marker can be used to select for the desired form of the gene. For example, a soybean breeder can use a DNA marker to identify plants that carry the form of the gene that gives resistance to a disease rather than the form that leads to susceptibility. With the recent release of the whole DNA sequence of the soybean genome it became possible to identify thousands of DNA markers called Simple Sequence Repeat or SSR markers across the 20 soybean chromosomes. ARS scientists at the Soybean Genomics and Improvement Laboratory in Beltsville, MD with collaborating ARS scientists at Ames, IA screened the DNA sequence of the 20 soybean chromosomes and the identified more than 33,000 SSR markers with a high probability of functioning well for use in DNA marker assisted soybean breeding and for the discovery of the positions of genes on the soybean chromosomes. A database called BARCSOYSSR_1.0 was created which contains the information required for the use of each of the more than 33,000 SSR DNA markers as well as the specific position of each marker on one of the 20 soybean chromosomes. This information is available on SoyBase (http://soybase.org), the USDA, ARS, Soybean Genome Database. The information in this database will be useful to soybean breeders and soybean geneticists to select useful DNA markers at any position on any one of the 20 soybean chromosomes to facilitate gene cloning or DNA marker assisted soybean breeding.


Review Publications
Schmutz, J., Cannon, S.B., Schlueter, J., Ma, J., Hyten, D.L., Song, Q., Mitros, T., Nelson, W., May, G.D., Gill, N., Peto, M.F., Shu, S., Goodstein, D., Thelen, J.J., Cheng, J., Sakurai, T., Umezawa, T., Shinozaki, K., Du, J., Bhattacharyya, M., Sandhu, D., Grant, D.M., Joshi, T., Libault, M., Zhang, X., Hguyen, H., Valliyodan, B., Xu, D., Futrell-Griggs, M., Abernathy, B., Hellsten, U., Berry, K., Grimwood, J., Yu, Y., Wing, R.A., Cregan, P.B., Stacey, G., Specht, J., Rokhsar, D., Shoemaker, R.C., Jackson, S. 2010. Genome Sequence of the Paleopolyploid Soybean (Glycine max (L.) Merr.). Nature. 463:178-183.

Hyten, D.L., Choi, I., Song, Q., Specht, J.E., Carter Jr, T.E., Shoemaker, R.C., Hwang, E., Matukumalli, L.K., Cregan, P.B. 2001. A high density integrated genetic linkage map of soybean and the development of a 1,536 Universal Soy Linkage Panel for QTL mapping. Crop Science. 50:960-968.

Hyten, D.L., Cannon, S.B., Song, Q., Weeks, N.T., Fickus, E.W., Shoemaker, R.C., Specht, J.E., May, G.D., Cregan, P.B. 2010. High-Throughput SNP Discovery through Deep Resequencing of a Reduced Representation Library to Anchor and Orient Scaffolds in the Soybean Whole Genome Sequence. Biomed Central (BMC) Genomics. 11:38.

Kim, K.S., Bellendir, S., Hudson, K. A., Hill, C.B., Hartman, G.L., Hyten, D., Hudson, M.E., Diers, B.W. 2010. Fine mapping the soybean aphid resistance gene Rag1 in soybean. Theoretical and Applied Genetics. 20(5):1063-1071.

Last Modified: 9/21/2014
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