"Nonrandom" DNA sequence analysis in bacteriophage M13 by the dideoxy chain-termination method.
"Nonrandom" DNA sequence analysis in bacteriophage M13 by the dideoxy chain-termination method.
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通过双脱氧链终止法对噬菌体 M13 进行“非随机”DNA 序列分析。
DOI：
10.1073/pnas.79.14.4298
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发表时间：
1982
影响因子：
11.1
通讯作者：
Surrey,S
中科院分区：
文献类型：
--
作者：
Poncz,M;Solowiejczyk,D;Ballantine,M;Schwartz,E;Surrey,S
我们描述了一种快速的“非随机”DNA序列分析程序，有助于确定DNA的大连续区域的核苷酸序列。该方法包括将感兴趣的限制性内切酶片段克隆到噬菌体M13中，然后从M13中靠近DNA插入的单个位点构建一系列核酸酶BAL-31缺失突变体。测定缺失突变体的大小是通过与含有总插入的M13衍生的互补单链探针杂交，然后进行核酸酶S1处理来完成的。使用位点特异性M13 DNA引物和双脱氧链终止法，分离和分析了越来越小的单链M13插入DNA。通过这种方式，DNA序列的分析以一种非随机的方式从总插入的一端进行到另一端，因为产生了一组可控的重叠缺失突变体。
We describe a rapid "nonrandom" DNA sequence analysis procedure that facilitates the nucleotide sequence determination of large contiguous regions of DNA. The method consists of cloning a restriction endonuclease fragment of interest into bacteriophage M13 followed by construction of a series of nuclease BAL-31 deletion mutants originating from a single site in M13 that is close to the DNA insert. Determination of the size of the deletion mutant is accomplished by hybridization to a complementary single-stranded probe derived from M13 containing that total insert followed by nuclease S1 treatment. Single-stranded M13-insert DNAs of progressively smaller sizes are isolated and analyzed by using a site-specific M13 DNA primer and the dideoxy chain-termination method. In this way, analysis of the DNA sequence proceeds from one end of the total insert to the other in a nonrandom fashion due to generation of a controlled overlapping set of deletion mutants.