普氏原羚粪便DNA提取方法的改进与比较

Examination and Comparison of Four Methods on DNA Extraction from the Faeces of Procapra praprzewalskii

为提高普氏原羚粪便DNA提取效果,在总结传统提取粪便DNA方法的基础上,设计了新的提取方法。该方法以NaCl替代TE溶解粪便样品,增加溶菌酶用量以减少蛋白酶K用量。经琼脂糖凝胶电泳和PCR扩增12SrRNA片段对改进方法与传统提取粪便DNA方法进行比较,发现用改进方法提取的DNA质和量均比传统方法好,且可满足一般分子生物学实验的需要。扩增的普氏原羚12SrRNA片段已经测序,并递交GenBank,登录号为EU247756。新方法不仅克服了传统提取法中DNA降解严重的问题,而且有效地解除了粪便DNA对Taq酶的抑制作用,为普氏原羚遗传多样性保护提供技术支持。

A new method was developed to extract DNA from the faeces of Procapra praprzewalskii on the base of concluding three old methods. 12SrRNA gene was amplified and sequenced successfully from faecal DNA, suggesting that the method of DNA extrac- tion was reliable and efficient. This method not only overcame previous barriers to molecular scatology, such as inhibitors, low quality ,degradation of DNA, but also was free of inhibitor to Taq. The 12SrRNA gene accession is EU247756. These results provide baseline data for the evaluation of the level of genetic variation in Procapra praprzewalskii, which will be important for the development of conservation strategies in future.