A METHOD FOR DIFFERENCE CLONING - GENE AMPLIFICATION FOLLOWING SUBTRACTIVE HYBRIDIZATION

被引：96

作者：

WIELAND, I ^{[1
]}

BOLGER, G ^{[1
]}

ASOULINE, G ^{[1
]}

WIGLER, M ^{[1
]}

机构：

[1] COLD SPRING HARBOR LAB,COLD SPRING HARBOR,NY 11724

来源：

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA | 1990年 / 87卷 / 07期

关键词：

cancer; infectious disease; inherited disease; polymerase chain reaction;

D O I：

10.1073/pnas.87.7.2720

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

07 ; 0710 ; 09 ;

摘要：

We describe a procedure for genomic difference cloning, a method for isolating sequences present in one genomic DNA population ('tester') that is absent in another ('driver'). By subtractive hybridization, a large excess of driver is used to remove sequences common to a biotinylated tester, enriching the 'target' sequences that are unique to the tester. After repeated subtractive hybridization cycles, tester is separated from driver by avidin/biotin affinity chromatography, and single-stranded target is amplified by the polymerase chain reaction, rendering it double-stranded and clonable. We model two situations: the gain of sequences that result from infection with a pathogen and the loss of sequences that result from a large hemizygous deletion. We obtain 100- to 700-fold enrichment of target sequences.

引用

页码：2720 / 2724

页数：5

共 28 条

[1] CHARACTERIZATION OF AN ACTIVATED HUMAN ROS GENE [J].