牦牛乳中DNA提取方法的建立与优化

Establishment and optimization of the DNA extraction from yak milk

摘要:

为加速牦牛重要基因的挖掘和功能鉴定,以牦牛乳作为试验材料,建立简便的牦牛基因组DNA提取方法.采用常规离心法,从牦牛乳中获得体细胞,进而提取基因组DNA.研究不同去除蛋白方法、不同DNA沉淀剂、不同贮藏条件对牦牛乳中DNA提取质量的影响.DNA质量通过PCR评价.结果表明:高盐法与酚/氯仿法去除蛋白效果相当.高盐法简便快速,且避免了酚、氯仿等有机试剂对人们健康的影响.异丙醇和无水乙醇的沉淀DNA效果相近,而乙二醇丁醚效果最差.牦牛乳贮藏条件对DNA质量影响很大,4℃下短时间保存或在-20℃下长期保存,均可获得理想的DNA.采用高盐法从牦牛乳中提取DNA可用于PCR特异性扩增等后续研究工作.

Abstract:

A reliable and rapid method for genomic DNA extraction was constructed here to accelerate the identification of important genes in yak.In this method,the yak milk was first centrifuged,and then somatic cells were obtained for DNA extraction.The influences of different methods of protein removal,DNA precipitants,and storage conditions of yak milk on DNA quality were discussed.The quality of DNA was detected using PCR.Experimental results indicated that both the salting-out and phenol-chloroform method were effective for the removal of proteins,however the salting-out method was more simple and rapid.The precipitants isopropyl alcohol and ethanol were similar to precipitate DNA,but the glycol ether was the worst precipitant in this study.The storage conditions of yak milk have remarkable influence on the quality of DNA,and when the yak milk was stored at 4 ℃ for short term,or-20 ℃ for long term,the desired DNA can be got.The DNA extracted from yak milk using the salting-out method can be used in the later genetic research.