Submitted to: International Cotton Genome Initiative Workshop
Publication Type: Proceedings
Publication Acceptance Date: 1/9/2002
Publication Date: 6/3/2002
Citation: YU, J., KOHEL, R.J., ZHANG, H., DONG, J., SUN, S., STEELE, N. TOWARD DEVELOPMENT OF A WHOLE-GENOME, BAC/BIBAC-BASED INTEGRATED PHYSICAL/GENETIC MAP OF THE COTTON GENOME USING THE UPLAND GENETIC STANDARD TM-1: BAC AND BIBAC LIBRARY CONSTRUCTION, SSR MARKER DEVELOPMENT, AND PHYSICAL/GENETICS. PROCEEDINGS OF INTERNATIONAL COTTON GENOME INITIATIVE WORKSHOP. 2002. V. 14. P. 32.
Technical Abstract: Integrative physical mapping is the centerpiece of and essential for advanced genomics research. Upland cotton (Gossypium hirsutum L.) genetic standard line TM-1 is used as the reference genotype to develop a whole-genome, BAC/BIBAC-based integrated physical/genetic map of the cotton genome. From the TM-1 line we have constructed two BAC libraries with HindIII and EcoRI, respectively, and one plant-transformation-competent binary BAC (BIBAC) library with BamHI. Three large-insert BAC and BIBAC libraries constructed with different restriction enzymes not only reduce the number of clones needed, but also enhance the coverage of a genome-wide physical map. TM-1 is used for this effort because from this line extensive genetic mutants and cytogenetic stocks were developed. In addition, TM-1 is one of the two parents of our permanent recombinant inbred (RI) mapping population used in the integrated genetic/physical mapping of the cotton genome. Furthermore, seed stock of TM-1 for the libraries was maintained at 54th selfing generation by single-seed decent to ensure the plant homogeneity and homozygosity that are essential to a successful assembly of BAC contig maps by fingerprint analysis that truly reflect the genome structure of the Upland cotton. The three TM-1 BAC/BIBAC libraries contain 158,000 large-insert clones, have an average insert size of 130, 152 and 154 kb, respectively and cover >10x of the haploid (AD)1 genomes. They are now being used to construct the whole-genome BAC/BIBAC-based integrated genetic/physical map of Upland cotton and other aspects of genomic research. To have a large number of portable SSR markers for the cotton genetic map and facilitate physical/genetic map integration, we have been developing SSR markers from the TM-1 BAC libraries. The BAC-derived SSR markers have many advantages over those developed from small-insert DNA clones. The SSR marker-containing BACs provide direct bridges to physical contig maps with genetic linkage maps. They will not only streamline high-resolution mapping and positional cloning of QTLs and genes of interest, but also lead to the development of many different kinds of DNA markers that are well suited for marker-assisted breeding. An initial set of 1,000 such SSR primer pairs have been developed from the TM-1/HindIII BAC library. Approximately 60% of these BAC-SSR primer pairs amplify only single fragments, which are very useful for integration of physical/genetic maps and identification of the A- or D-subgenome specific BAC contigs. High-density filters and pools of the TM-1/HindIII BAC clones have been distributed to the international cotton research community for specific genomic studies. TM-1 BAC-derived SSR markers will be distributed to the cotton research community once we determine the value of polymorphism information content (PIC) on each SSR marker with a diverse cotton panel. The TM-1 x 3-79 genetic map has approximately 1,000 DNA markers and it is now being augmented with the 1,000 BAC-derived SSR markers and other 1,000 SSR markers currently funded by the Cotton Incorporated. Integration of genetic and physical maps with a large number of portable SSR markers on the BAC contigs will certainly benefit the cotton genome research including molecular breeding, germplasm evaluation, and functional analysis of thousands of cotton genes.