Control of pH during plasmid preparation by alkaline lysis of Escherichia coli

被引：8

作者：

Cloninger, Cheri ^{[1
]}

Felton, Marilyn ^{[1
]}

Paul, Bonnie ^{[1
]}

Hirakawa, Yasuko ^{[1
]}

Metzenberg, Stan ^{[1
]}

机构：

[1] Calif State Univ Northridge, Dept Biol, Northridge, CA 91330 USA

来源：

ANALYTICAL BIOCHEMISTRY | 2008年 / 378卷 / 02期

关键词：

D O I：

10.1016/j.ab.2008.04.017

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

Alkaline lysis of Escherichia coli is usually the method of choice for plasmid preparation, but "ghost bands" of denatured supercoiled DNA can result if the pH is too high or the period of lysis is too long. By replacing the usual sodium hydroxide lysis solution with an arginine buffer prepared in the range of pH 11.4 to 12.0, we were able to stabilize the pH during lysis and obtain plasmid that is suitably pure for restriction digestion and DNA sequencing. (C) 2008 Elsevier Inc. All rights reserved.

引用

页码：224 / 225

页数：2

共 10 条

[1]

BIRNBOIM HC, 1979, NUCLEIC ACIDS RES, V7, P1513

[2] Rapid isolation of high quality, multimeric plasmid DNA using zwitterionic detergent [J].

Chowdhury, EH ;

Akaike, T .

JOURNAL OF BIOTECHNOLOGY, 2005, 119 (04) :343-347

[3] ISOLATION OF COVALENTLY CLOSED CIRCULAR DNA OF HIGH MOLECULAR-WEIGHT FROM BACTERIA [J].

CURRIER, TC ;

NESTER, EW .

ANALYTICAL BIOCHEMISTRY, 1976, 76 (02) :431-441

[4] METHODS AND REAGENTS - GHOST PLASMID OF PBLUESCRIPT [J].

HENGEN, PN .

TRENDS IN BIOCHEMICAL SCIENCES, 1994, 19 (03) :139-140

[5] Methods and reagents - Eliminating ghost bands from plasmid preps [J].

Hengen, PN .

TRENDS IN BIOCHEMICAL SCIENCES, 1996, 21 (11) :441-442

[6]

LIDE DR, 1990, CRC HDB CHEM PHYSICS, P7

[7]

*QIAG, 2005, QIAG PLASM PUR HDB

[8] THE ENRICHMENT OF PLASMID DNAS, IN BACTERIAL-CELL LYSATES, USING AN ALKALINE-PH PROCEDURE THAT DOES NOT PERMANENTLY DENATURE THEM [J].

RANHAND, JM .

PREPARATIVE BIOCHEMISTRY, 1985, 15 (03) :121-131

[9] Identification and eradication of a denatured DNA isolated during alkaline lysis-based plasmid purification procedures [J].

Sayers, JR ;

Evans, D ;

Thomson, JB .

ANALYTICAL BIOCHEMISTRY, 1996, 241 (02) :186-189

[10] PHYSICAL AND TOPOLOGICAL PROPERTIES OF CIRCULAR DNA [J].

VINOGRAD, J ;

LEBOWITZ, J .

JOURNAL OF GENERAL PHYSIOLOGY, 1966, 49 (6P2) :103-&

← 1 →