小鼠冷冻胚胎和精子SNP遗传鉴定方法的建立

Establishment of a SNP genetic identification method for frozen embryos and sperm of inbred mice

目的建立小鼠冷冻胚胎和精子SNP(single nucleotide polymorphism)分型方法,用于冷冻胚胎和精子快速遗传鉴定方案。方法以中科院上海实验动物中心(国家啮齿类实验动物种子中心上海分中心)提供的小鼠冷冻胚胎和精子为样本,采用全基因组扩增技术和PCR-LDR分型技术建立小鼠冷冻物SNP遗传鉴定方法。结果全基因组扩增技术能大幅度增加冷冻胚胎样本的DNA总量;PCR-LDR分型方法适用于小鼠全基因组45个SNPs的分型;分型确定C57BL/6,BALB/c,FVB/NJ等胚胎和精子各10种近交系,SNP位点信息与测序结果一致;小鼠冷冻胚胎个数与SNPs检出个数成正比,当胚胎数达到12以上时SNP检出率100%。结论实现近交系小鼠冷冻胚胎和精子快速SNP基因分型及遗传质量鉴定。

Objective To establish a rapid SNP（ single-nucleotide polymorphism） genetic identification method for the frozen samples,such as frozen embryos and sperm of inbred mice. Methods In this study,the frozen embryos and sperm of inbred mice were provided by Shanghai Lab. Animal Research Center. Whole genome amplification and PCR-LDR genotyping system were used to get the rich DNA sample. Forty-five SNP were genotyped by multiple polymerase chain reaction and ligase detection reaction（ PCR-LDR）. Results The electrophoresis results showed that the whole genome amplification technique could highly increase the total DNA of frozen embryos. PCR-LDR typing method was suitable for the mouse genome typing of 45 SNPs. Ten strains of inbred frozen embryos and sperms of C57 BL /6,BALB / c,FVB / NJ mice were genotyping identified,and their SNP loci data obtained by PCR-LDR were as the same as those of database. The number of frozen mouse embryos was proportional to the number of SNPs detected,and when the embryo number reached more than 12,the detection rate of SNP was 100%. Conclusions This method can be used to the genetic quality identification,and rapidly identify the inbreed frozen mouse embryos and sperms.