Molecular Markers Of SSRs And SNPs Derived From ESTs And Their Application

The exponentially increased expressed sequence tags (ESTs) in public databases provide a potential resource for genetic analysis. Over 85Mb publicly available ESTs data of four agronomically important crops were used to search for the types and frequencies of simple sequence repeats (SSRs) with motif length of l-6bp long. The frequency of EST-SSRs was one every 11.81kb in rice, 17.42kb in wheat, 23.80kb in soybean, and 28.32kb in maize, respectively. Trinucleotide repeats were the most abundant SSR types with up to 47 trinucleotide repeat units in 100kb ESTs. Compared with the dicot, the monocots preferred GC-rich tri- and hexa- motifs very much, especially in rice where 23.6 CCG motif occurred in 100kb ESTs. 597 EST-SSR primer pairs were designed for wheat, of which, 478 primer pairs had successful PCR amplifications. The percentage of polymorphism was up to 41.8% among wheat varieties, which could be used to construct transcriptional map of wheat. Cross-species amplification was detected. Among the 478 functional primers from wheat, 255 EST-SSRs amplified fragments in rice, maize and soybean too, indicating high degree of transferability between species, which facilitated comparative mapping and homologous gene cloning.Four hundred and senventy-eight EST-SSR working markers were screened among three parents of mapping populations (W-7984xOpata 85, WOpop; LumaixHanxuan, LHpop; WenmaixShanhongmai, WSpop). The number of polymorphic EST-SSR primer pairs between parents of WOpop, LHpop and WSpop was 92, 58 and 29 respectively. A total of 101 EST-SSR loci amplified from 88 primer sets were distributed onto the 20 chromosomes of the reference maps (no marker on chromosome 4B). The 101 mapped EST-SSR loci added to the existing 450 microsatellite loci previously mapped in bread wheat. 74 of the 101 loci showed significant similarities to known genes, which included 24 genes involved in metabolism, 4 in cellular structures, 9 in stress resistance, 12 in transcription, 2 in development, 2 transporters and 21 storage proteins. Besides gliadin and glutenin, most of the genes with putative function (53) were mapped for the first time by EST-SSR markers in bread wheat. Sequence alignment of mapped wheat EST-SSR loci allowed tentative assignment of functionality to the other members of grasses family. Colinearity combined with homology information offers an attractive approach to comparative genomics.SNPs are the most abundant resource of genetic variation among individuals of a species. SNP investigation on plants is still at the beginning stage, especially for polyploidy plant genomes, such as hexaploid wheat. An open international consortium of institutions (public and private) (http://wheat.pw.usda.gov/ITMl/WheatSNP/) is attempting to mine the contigs in a coordinated way, pooling information on validated SNPs. As a member of Wheat SNP Development, we are responsible for validating SNPs from 20 contigs allocated. It suggested that ABI Prism?SNaPshot?is powerful for validating the SNPs.