用酚-氯仿直接从克隆中提取质粒DNA

A Method of Direct Extraction of Plasmid DNA with Phenol-chloroform from Clone

目的:对用酚-氯仿直接从克隆中提取质粒DNA的方法进行了研究。方法:采用不同次数的酚-氯仿抽提,在不同RNaseA作用温度及作用时间下提取了三种质粒。结果:用酚-氯仿抽提2次,RNase A45℃作用8min,质粒纯度(OD260/OD280)为1.85左右、产量大于0.8μg/克隆。结论:本法不但快速,且所提质粒均能满足后续实验如酶切鉴定、测序等方面的需要。对于大批量重组克隆的筛选尤为适合。

Objective：Study on extraction of plasmid DNA with phenol-chloroform from clone.Methods：Adopt different extraction times with phenol-chloroform,different acting temperature and acting time of RNaseA.we extracted three kinds of plasmid DNA.Results：extraction twice with phenol-chloroform,acting time of RNaseA was 8 minutes in 45 ℃.The agarose electrophoresis showed that the quality of plasmid DNA was about OD260/OD280=1.85,and quantity was more than 0.8μg/clone.Conclusion：This method is not only rapid, but also can satisfy the demands of analysis of recombinant plasmids and sequence measure. This method is adapt to sieving of a large amount of clone specially.