家蚕胚胎细胞系的DNA指纹图谱分析

在建立可靠的家蚕细胞系基因组DNA制备和PCR扩增技术体系的基础上,筛选具有稳定多态性位点的RAPD和ISSR引物,建立家蚕细胞系基因组DNA的ISSR和RAPD分子标记技术体系,检测家蚕细胞系的DNA分子标记多态性,构建细胞系的DNA指纹图谱。筛选出了26个ISSR引物和43个RAPD引物,通过PCR扩增在家蚕胚胎细胞系和传代昆虫细胞系等9个样品中分别获得了797条和1205条多态性条带,多态性达到89．9%和76．6%,不同细胞系的DNA多态性有较大差异,三个家蚕胚胎细胞系具有各自特有的DNA标记。测定了9个样品间的Nei's相似系数和遗传距离,构建了系统发育树,结果表明本实验室建立的3个家蚕胚胎细胞系和家蚕“夏芳×秋白”聚为一簇,亲缘关系较近,而来自不同物种的五个传代昆虫细胞系聚为一簇,它们之间的遗传距离比3个家蚕胚胎细胞系之间的遗传距离更小。

DNA FINGERPRINTING ANALYSIS OF SILKWORM EMBRYO CELL LINES

DNA extraction and the polymerase chain reaction (PCR) were used on DNA genomes study of cell lines of Bombyx mori.DNA polymorphic marker analysis was conducted and DNA fingerprint of cell lines of Bombyx mori.was carried out using ISSR and RAPD. Primers that can reliably find polymorphic bands were screened out.26 ISSR primers were selected from them any available,and 797 polymorphic bands were abtained through PCR amplification in 9 samples,including 3 embryo cell lines of Bombyx mori (BmE-SWU1,BmE- SWU2,BmE-SWU3),5 passage cell lines (BmE,BmN,Sf9,Sf21,Hi5)and the embryos from which BmE-SWU1 originated.The ration of polymorphic bands was 89.9%.43 RAPD primers were selected out through PCR amplification,and 1205 polymorphic bands were obtained in 9 samples. The ration of polymorphic bands was 76.6%.There were many DNA polymorphic bands differences in the cell lines of Bombyx mori.The special DNA markers of the 3 embryo cell lines were found respectively.The similarity index Nei and genetic distance of the 9 samples were calculated and the phylogeny tree of 9 samples was constructed by UPGMA.Results showed that 2 groups were divided,one group including the 3 embryo cell lines and the embryo of XQ has close relative.Another group constructed by five insect cell lines came from different species,their genetic distance was closer than the 3 embryo cell lines.