一种用质粒DNA转化大肠杆菌感受态细胞的实用操作技巧  

Doohickey for the Standard Transformation Protocol of Plasmids DNA Transfer into Competent Escherichia coil

作　　者：陈洪栋[1] 董文博 李红民[1] 

机构地区：[1]西部资源生物与现代生物技术教育部重点实验室植物生物技术研究室西北大学生命科学学院,西安710069

出　　处：《生物技术通报》2009年第S1期297-299,共3页Biotechnology Bulletin

基　　金：西安市科技局科技攻关项目(GG06078)

摘　　要：目的是建立一种简化、实用的用质粒DNA转化大肠杆菌的操作方法。采用氯化钙法制备大肠杆菌感受态细胞。以质粒pUC18,pCSN44,pAN52-1Not,pETts,pANth和植物双元表达载体pCAMBIA1301分别转化用于质粒扩增与保存的常用大肠杆菌菌株Top10和DH5α以及用于原核表达的常用大肠杆菌菌株BL21(DE3)和TB1。质粒与感受态细胞的混合液置冰上作用一定时间后,直接涂布含有筛选抗生素的LB平板,于37℃培养12～16h。结果表明,用不同大小的质粒DNA转化不同的大肠杆菌菌株,都可以获得满足实验要求,转化效率可高达103～4阳性克隆/μg。该方法较标准的转化流程更加简便、省时、实用。The aim of this study was to introduce a doohickey for the transformation of competent E.coli cells with plasmid DNA. The competent cells of Escherichia coli strains were previously made using the calcium chloride method.The plasmids,pUC18,pCSN44, pAN52-lNot,pETts,pANth and pCAMBIA1301,were employed to transform the different E.coli strains Top10,DH5α,BL21(DE3) and TB1.The mixture of plasmid and competent cell was laid on ice for definite time before being cultured at 37℃degree on selected LB plate with relevant antibiotic for 12～t6h.Perfect transformation rate up to 103～4 colonies/μg plasmid DNA was obtained.

关 键 词：ESCHERICHIA COLI 转化方法 质粒 

分 类 号：Q78[生物学—分子生物学]