编码MspI限制修饰系统的基因的克隆与特性分析
Cloning and characterization of the genes encoding the MspI restriction modification system.
作者信息
Lin P M, Lee C H, Roberts R J
机构信息
Cold Spring Harbor Laboratory, NY 11724.
出版信息
Nucleic Acids Res. 1989 Apr 25;17(8):3001-11. doi: 10.1093/nar/17.8.3001.
Abstract
The genes encoding the MspI restriction modification system, which recognizes the sequence 5' CCGG, have been cloned into pUC9. Selection was based on expression of the cloned methylase gene which renders plasmid DNA insensitive to MspI cleavage in vitro. Initially, an insert of 15 kb was obtained which, upon subcloning, yielded a 3 kb EcoRI to HindIII insert, carrying the genes for both the methylase and the restriction enzyme. This insert has been sequenced. Based upon the sequence, together with appropriate subclones, it is shown that the two genes are transcribed divergently with the methylase gene encoding a polypeptide of 418 amino acids, while the restriction enzyme is composed of 262 amino acids. Comparison of the sequence of the MspI methylase with other cytosine methylases shows a striking degree of similarity. Especially noteworthy is the high degree of similarity with the HhaI and EcoRII methylases.
摘要
编码识别序列5' CCGG的MspI限制修饰系统的基因已被克隆到pUC9中。选择是基于克隆的甲基化酶基因的表达，该基因使质粒DNA在体外对MspI切割不敏感。最初，获得了一个15 kb的插入片段，该片段在亚克隆后产生了一个3 kb的EcoRI至HindIII插入片段，携带甲基化酶和限制酶的基因。这个插入片段已经测序。根据该序列以及合适的亚克隆，结果表明这两个基因是反向转录的，甲基化酶基因编码一个418个氨基酸的多肽，而限制酶由262个氨基酸组成。MspI甲基化酶与其他胞嘧啶甲基化酶的序列比较显示出惊人的相似程度。特别值得注意的是与HhaI和EcoRII甲基化酶的高度相似性。
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