Location: Crop Germplasm Research2012 Annual Report
1a. Objectives (from AD-416):
Objective 1: Identify a core set of molecular markers tailored for systematic characterization of the genetic diversity within and among Gossypium germplasm accessions that will be maintained under the sister project 6202-21000-032-00D. Objective 2: Maintain and enhance CottonDB as a user-friendly tool for the cotton research community. Sub-objective 2.A: Maintain and enhance CottonDB, including development of user friendly public interfaces. Sub-objective 2.B: Develop bioinformatic software and tools to assist both users and curators of CottonDB. Objective 3: Collaborate with other public sector researchers to construct and integrate physical and genetic maps of G. hirsutum. Sub-objective 3.A: Develop cotton genetic maps that contain PCR-based DNA markers. Sub-objective 3.B: Develop cotton physical maps that contain large-insert BAC clones. Sub-objective 3.C: Integrate cotton genetic and physical maps with EST unigene information. Objective 4: Identify key genes and genomic regions of cotton for use in developing cotton germplasm resources that exhibit desirable/improved agronomic and fiber traits. Sub-objective 4.A: Apply genomic and bioinformatic tools to identify and characterize QTLs or alleles from cotton genetic resources, maintained under the sister project 6202-21000-032-00D, that govern key agronomic or fiber traits. Sub-objective 4.B: Apply the preceding information to identify superior parents for developing breeding populations with novel sources of variability for traits of interest. Sub-objective 4.C: Recombine and select the preceding breeding populations to accumulate desirable QTLs and alleles in enhanced cotton breeding lines.
1b. Approach (from AD-416):
To develop a portable core set of markers for cotton (Objective 1), new SSR and SNP markers will be developed from cotton BAC libraries and other genomic DNA templates. From the markers created, a core set of 208 markers will be carefully selected from the saturated genome map of tetraploid cotton (TM-1 x 3-79) with 8 markers from each of 26 chromosomes. Each of these core markers will have a high polymorphism information content (PIC) value to be determined on a standardized core germplasm panel consisting of 12 diverse Gossypium genotypes. These markers will be evenly distributed on the cotton genome, with every chromosome arm having 4 core markers at approximately 15-cM intervals. Data from marker development will be stored and made available in the CottonDB database. CottonDB, a tool for the research community, will be enhanced through continued migration of its information content to a relational structure, improved display pages, and direct record-to-record links between internet databases to integrate information into a larger virtual database (Objective 2). To enrich the delivered content and streamline users' searches for specific information, work will integrate related data from multiple databases. Solutions developed by other genome databases will be adapted and implemented to this project's databases where appropriate. To construct and integrate physical and genetic maps, genetic mapping of TM-1 BAC-derived and other markers will be conducted using the TM-1 x 3-79 RI population. Diagnostic DNA markers will be identified that are capable of detecting polymorphism in intraspecific populations, and these markers will be used to genotype the entire TM-1 x 3-79 RI mapping population. A score matrix will be generated from the genotyping experiments and merged with the existing mapping database to perform linkage analysis via MapMaker and/or JOINMAP software programs. Recombination frequencies will be converted into map distances (cM). Approximately 500 SSR and 500 SNP markers will be added to the existing genetic map that contains 1,200 SSR markers to obtain an average resolution of 1-2 cM per marker. Integration of cotton genetic and physical maps will be achieved by anchoring framework genetic markers to TM-1 BAC contigs, and locating BAC-derived markers to the TM-1 x 3-79 RI map (Objective 3). Comparisons of genetic and physical map tools (CMap and IntegratedMap) will allow for consolidation of all structural and physical genomic information. In order to utilize the growing numbers of QTLs reported in cotton, work will validate those QTL by aligning genomic locations and comparing genetic effects (Objective 4). Information for QTLs of interest will be related among comparable studies in cotton and will be obtained from a variety of sources, including published accounts and database records. Once specific chromosomal regions containing genes that make a significant contribution to the expression of a complex phenotype of interest are identified, fine-mapping of the most promising genomic regions will be used to identify polymorphisms in coding and/or regulatory regions.
3. Progress Report:
During FY 2012, a simplified method for studying the genetic characteristics of cotton was developed and tested in collaborative work with Texas A&M University. The method known as genotyping-by-sequencing (GBS) was simplified and used to discover new molecular markers known as SNPs. This simplified GBS approach provides a powerful tool for developing informative markers in cotton species that as yet have no sequenced genome. It also generates a valuable resource that is useful to whole genome sequencing projects. Also in FY 2012, significant progress was made in sequencing the wild D-genome cotton species G. raimondii in collaboration with international cooperators. The ancestor of G. raimondii is considered to be one of the parents of the modern economically important fiber-producing cotton species G. hirsutum and G. barbadense. Detailed structural, functional, and comparative analyses of the DNA sequence data revealed previously unknown genome complexity and identified important genes of interest. The DNA sequence developed by this work provides cotton researchers with a highly useful tool and will lead to an eventual sequence map of the economically important tetraploid cotton genomes. The merger of CottonDB (http://www.cottondb.org/) with Cotton Marker Database (http://www.cottonmarker.org/) was completed in FY 2012. The resulting database, known as CottonGen (http://www.cottongen.org/), is managed by Washington State University. The successful database merger has resulted in a significantly improved database, and has freed up fiscal resources for use by this project; the project provides limited support to CottonGen through a Specific Cooperative Agreement with Cotton Incorporated. A project scientist serves on the CottonGen Steering Committee.
1. High-density genetic map of cotton. A high-density genetic map of the cotton genome is necessary for quick, high efficiency molecular genetics techniques to be applied to cotton improvement. ARS scientists at College Station, Texas, working with collaborators throughout the U.S., developed a high density map covering all 26 cotton chromosomes with 2,527 molecular markers. This cotton genetic map with DNA markers provides useful resources for germplasm characterization, gene discovery, and molecular breeding. The map will ultimately facilitate development of better cotton varieties for U.S. farmers.
2. Core molecular marker set to define genetic diversity of cotton. Although molecular markers have been used to investigate the diversity of various groups and collections of cotton, the use of non-standardized, non-uniform marker sets does not allow us to look at the diversity of cotton across all groups and collections. ARS scientists at College Station, Texas, developed a set of 208 markers on the basis of their variability within cotton, and their being evenly distributed across the 26 chromosomes of cotton. Of the 208 markers, an initial set of 105 have been selected as a core set and they have been used to characterize the genetic diversity of the U.S. Cotton Germplasm Collection. These 105 core DNA markers are being requested by many in the national and international cotton research communities and should provide a common basis for systematic characterization of cotton germplasm collections worldwide.
Yu, J., Kohel, R.J., Fang, D.D., Cho, J., Van Deynze, A., Ulloa, M., Hoffman, S.M., Pepper, A.E., Stelly, D.M., Jenkins, J.N., Saha, S., Kumpatla, S.P., Shah, M.R., Hugie, W.V., Percy, R.G. 2012. A high-density simple sequence repeat and single nucleotide polymorphism genetic map of the tetraploid cotton genome. Genes, Genomes, Genetics. 2:43-58.