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采用热休克法将质粒DNA转化到大肠杆菌中。

Transformation of plasmid DNA into E. coli using the heat shock method.

作者信息

Froger Alexandrine, Hall James E

机构信息

Department of Physiology and Biophysics, University of California, Irvine, USA.

出版信息

J Vis Exp. 2007(6):253. doi: 10.3791/253. Epub 2007 Aug 1.

Abstract

Transformation of plasmid DNA into E. coli using the heat shock method is a basic technique of molecular biology. It consists of inserting a foreign plasmid or ligation product into bacteria. This video protocol describes the traditional method of transformation using commercially available chemically competent bacteria from Genlantis. After a short incubation in ice, a mixture of chemically competent bacteria and DNA is placed at 42 degrees C for 45 seconds (heat shock) and then placed back in ice. SOC media is added and the transformed cells are incubated at 37 degrees C for 30 min with agitation. To be assured of isolating colonies irrespective of transformation efficiency, two quantities of transformed bacteria are plated. This traditional protocol can be used successfully to transform most commercially available competent bacteria. The turbocells from Genlantis can also be used in a novel 3-minute transformation protocol, described in the instruction manual.

摘要

使用热休克法将质粒DNA转化到大肠杆菌中是分子生物学的一项基本技术。它包括将外源质粒或连接产物导入细菌。本视频方案描述了使用Genlantis公司市售化学感受态细菌进行转化的传统方法。在冰上短暂孵育后,将化学感受态细菌和DNA的混合物置于42℃ 45秒(热休克),然后放回冰上。加入SOC培养基,将转化后的细胞在37℃振荡孵育30分钟。为确保无论转化效率如何都能分离到菌落,接种两份转化后的细菌。这个传统方案可以成功用于转化大多数市售的感受态细菌。Genlantis公司的TurboCells也可用于说明书中描述的新型3分钟转化方案。

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