一种在宽尺寸范围内快速纯化共价闭合环状质粒DNA的改进方法。

An improved method for rapid purification of covalently closed circular plasmid DNA over a wide size range.

作者信息

Azad A K, Coote J G, Parton R

机构信息

Department of Microbiology, University of Glasgow, UK.

出版信息

Lett Appl Microbiol. 1992 Jun;14(6):250-4. doi: 10.1111/j.1472-765x.1992.tb00698.x.

DOI:10.1111/j.1472-765x.1992.tb00698.x

PMID:1368369

Abstract

An improved method has been developed for the large-scale purification of covalently closed circular (CCC) plasmid DNA molecules of sizes ranging from 4.3 to 73 kb. This protocol uses an alkaline-lysis procedure followed by acid-phenol extraction but with several modifications to previously reported methods. The principal modification is the replacement of NaCl by MgCl2 in the extraction buffer to improve yield and to remove chromosomal and other non-CCC plasmid DNA. Plasmid DNA can be purified in less than 1 h and used successfully in restriction enzyme analysis and cloning experiments.

摘要

已经开发出一种改进方法，用于大规模纯化大小在4.3至73 kb之间的共价闭合环状（CCC）质粒DNA分子。该方案采用碱裂解程序，随后进行酸酚抽提，但对先前报道的方法进行了若干修改。主要修改是在抽提缓冲液中用MgCl2代替NaCl，以提高产量并去除染色体和其他非CCC质粒DNA。质粒DNA可在1小时内纯化出来，并成功用于限制性酶切分析和克隆实验。