Development And Application Of Microsatellite Markers And Caps Markers To Analyze Genetic Diversity In Silkworm, Bombyx Mori

In this paper, we constructed a 7 kb-insert genomic library from the silkworm strain 'P50'. The screening of the genomic library by the conventional hybridization method led to the isolation of 38 microsatellites harbouring clones. Microsatellite polymorphism was assayed in 34 diverse silkworm strains. Of the 748 PCRs carried out, all but four produced amplified products of the expected size. 22 microsatellites were polymorphic, showing 2–16 alleles per locus (mean 8.2). The repeat length did not show any relationship with the degree of polymorphism in the present study. Heterozygosity ranged from 0.161-0.922 (mean 0.746). PIC ranged from 0.164–0.922 (mean 0.746). The result of UPGMA analysis is up to the geographical distribution of silkworm. Hence, microsatellites, ranked according to their information content, are recommended as markers of choice for silkworm fingerprinting and suggestions are provided for interpreting band profiles and the correct sizing of alleles.A silkgland DNA extraction method was developed for rapid identification of silkworm genotype. The method was applied to 6 silkworm strains. This method will be useful for genotypic selection that requires rapid screening of large populations by PCR and AFLP analysis. The method includes two major steps (i) grinding of a pair of silkglands with a roller in 1.5 ml tube and then treatment by proteinase K in an SDS extraction buffer, and (ii) precipitating protein with NaAC. About 200 μg of DNA per pair of silkglands of silkworm was obtained and more than 100 samples could be handled by one person in a day. 129 DNA sequences about silkworm were downloaded form genbank on internet. 98 primer pairs were designed from these sequences and 13 of them were polymorphism between 'Jingsong' and 'Lan10'. 2 primer pairs were found polymorphism between 'P50' and 'C108'. It will be the main way to develop molecular markers after the genomic sequence of silkworm was reported.