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球形红假单胞菌中一种位点特异性限制性内切酶的特性分析。

Characterization of a site-specific restriction endonuclease from Rhodopseudomonas sphaeroides.

作者信息

Lynn S P, Cohen L K, Gardner J F, Kaplan S

出版信息

J Bacteriol. 1979 May;138(2):505-9. doi: 10.1128/jb.138.2.505-509.1979.

DOI:10.1128/jb.138.2.505-509.1979
PMID:312287
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC218205/
Abstract

A type II restriction endonuclease, RshI, has been partially purified from photoheterotrophically grown Rhodopseudomonas sphaeroides strain 2.4.1. The enzyme preparation, after a single DE-52 column fractionation, is free of 5' exonuclease and phosphatase activities but contains a trace of 3' exonuclease activity. Based upon deoxyribonucleic acid (DNA) sequencing data in the vicinity of the enzyme-promoted cleavage of pBR322 DNA, we have concluded that RshI probably recognizes the palinodromic hexanucleotide sequence 5'-CGATCG-3' and cleaves between the T and C. lambda cI857 DNA contains three RshI sites, two of which lie in the replaceable region. The plasmid pBR322, which carries resistances to ampicillin and tetracycline, contains a single RshI site in the ampicillin resistance determinant. Insertion of DNA into the RshI site of pBR322 results in loss of ampicillin resistance but retention of tetracycline resistance, thereby providing a convenient screening procedure for recombinant plasmids.

摘要

一种II型限制性内切酶RshI已从光异养生长的球形红假单胞菌2.4.1菌株中部分纯化出来。该酶制剂经单一DE - 52柱分级分离后，无5'外切核酸酶和磷酸酶活性，但含有微量的3'外切核酸酶活性。根据pBR322 DNA酶促切割附近的脱氧核糖核酸（DNA）测序数据，我们得出结论，RshI可能识别回文六核苷酸序列5'-CGATCG-3'，并在T和C之间切割。λcI857 DNA含有三个RshI位点，其中两个位于可替换区域。携带氨苄青霉素和四环素抗性的质粒pBR322在氨苄青霉素抗性决定簇中含有一个单一的RshI位点。将DNA插入pBR322的RshI位点会导致氨苄青霉素抗性丧失，但四环素抗性保留，从而为重组质粒提供了一种方便的筛选方法。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ab69/218205/f27628126b2c/jbacter00282-0228-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ab69/218205/c22b6b4ea620/jbacter00282-0227-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ab69/218205/c087662070c2/jbacter00282-0228-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ab69/218205/f27628126b2c/jbacter00282-0228-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ab69/218205/c22b6b4ea620/jbacter00282-0227-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ab69/218205/c087662070c2/jbacter00282-0228-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ab69/218205/f27628126b2c/jbacter00282-0228-b.jpg

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Specific fragments of phi X174 deoxyribonucleic acid produced by a restriction enzyme from Haemophilus aegyptius, endonuclease Z.由埃及嗜血杆菌的一种限制酶——核酸内切酶Z产生的φX174脱氧核糖核酸的特定片段。
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