Development of RFLP markers from silkworm (Bombyx mori)

To construct RFLP linkage map of silkworm (Bombyx mori), as a preliminary step, we assessed the efficiency for single or low copy sequences from silkworm genomic DNA. A genomic DNA plasmid library was constructed using EcoRI-digested 0.3 to 5 kb DNA fragments of II-111 strain. As results of two step screening (reverse colony hybridization and reverse southern hybridization) using a total genomic DNA as a probe, about 6% (70/1,200) of clones examined was estimated single or low copy sequences. At least 80% of total clones contained middle or high repetitive sequences. These results indicate that B. mori genome has a lot of high repetitive sequences. However, analysis for its precise copy number in the genome remains to be elucidated. The obtained 70 single-sequences will be useful molecular markers to study silkworm genomic analysis in the near future, although it has low efficiency of single copy sequence for constructing linkage map.