"Improvement of linkage analysis in the silkworm, Bombyx mori, by using cDNA clones' RFLP."
2002
"Hara, W. (National Inst. of Agrobiological Sciences, Tsukuba, Ibaraki (Japan)) | Kosegawa, E. | Mase, K. | Kadono Okuda, K. "
"To make a molecular map of the silkworm, Bombyx mori, a high-yield and high-quality DNA was prepared from new BF1individuals, cDNA clones were prepared from a new library, and the scanning linkage analysis (SLA) was improved. To improve the yield and quality, 3- or 4-day-old 5th instar larvae were dissected and the gut was eliminated to avoid contaminating the mulberry leaf DNA. Hemolymph was removed to reduce the starting volume. High-quality DNA was purified by ultra-centrifugation in the CsCI gradient, and the yield exceeded 250 micro g per larva. A new cDNA library was constructed from 3-day-old embryonic mRNA using a plasmid as a vector. Preparing transformed bacteria and plasmid DNA from the cDNA library was easy, but the number of clones from the library was rather small compared to a previous library constructed using a phage as a vector. This was probably due to the low transformation efficiency of the plasmid vector. Using the newly prepared 15 BF1 individual DNA samples, the previously used 29 clones showed the same 9 linkage groups and 8 independent clones. Using these previous 29 clones and 11 new ones, 14 linkage groups and 6 independent clones covering 20 of 28 linkage groups were established. National Institute of Agrobiological Sciences, Tsukuba, Ibaraki 305-8634, Japan."
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