DNA分子标记检测家蚕苏菊、明虎品种和纯度的研究

Study on the Identification Method of Bombyx Variety and Purity of Suju and Minghu with DNA Molecular Marker

利用单粒卵微量DNA抽提法,从江苏省家蚕主要品种苏菊、明虎及其一代杂交种蚕卵快速提取出基因组DNA,用筛选出的 RAPD随机引物S77稳定扩增出苏菊、明虎的DNA特异性片段RS和RM,克隆和测序后大小分别为1694bp和1303bp,使用Blastn程序在NCBI 数据库检索发现,Rs的nt30-429与一个家蚕WGS(whole genome shotgun sequence)(dbj|BAAB01119085．1,contis504766)的nt472-874有92％的同源性,其中存在11个空缺位点(Gaps);RM的nt1070-1301与另一个家蚕WGS(dbj|BAAB01141578．1,contig554782)的nt262-31有91％的碱基相同,其中存在2个Gaps。进一步根据RS和RM序列合成SCAR标记引物PS-1(FP:5’ttccccccagtacgcaataac3’,RP:5’ ttccccccagacgtcacgtact3’)和PM-1(FP:5’ttcccccagtgatggaatttacg3’,RP:5’ttccccccagtccaatgttaca3’),能够准确、快速地鉴别所标记的苏菊、明虎及其F1蚕品种。

The genome DNA of Bombyx silkworm was extracted from single blue-stage-egg of mainly silkworm varieties in JiangSu province of Suju, Minghu and their Flwith rapid method. The specific bands RS-1690 and RM-1300 of RAPD were amplify traced each from of Suju and Minghu variety, with selected random primer of S77. The sequences of bands RS and RM were 1694bp and 1303bp respectively after cloned and sequenced. Then analyze the sequences by courtesy of database on-line with soft of Blastn. The results showed the nt30-429 of RS and the nt472-874 of the WGS（ whole genome shotgun sequence） （ dbj ｜ BAAB01119085.1 ｜ Bombyx mori DNA,contig504766） has 92% consistency, which include 11 gap sequences. Accordingly, 91% of nt262-31 of one of WGS （ dbj ｜ BAAB01141578.1 ｜ Bombyx rnori DNA,contig554782）shows the same sequence as the nt1070-1301 of RM, which include 2 gaps. Further, transform the RAPD mark of RS and RM to SCAR mark PS-I （ FP ：5＇ttccccccagtacgcaataac3＇, RP ：5＇ttccccccagacgtcacgtact3＇） and PM-1 （ FP：5＇ttcccccagtgatggaatttacg3＇, RP： 5＇ttccccecagtccaatgttaca3＇）. Thus, Suju, Minghu and their F1 were identified accu- rately and fast, with the synthetic specific primers of PS-1 and PM-1.