CLONING, OVEREXPRESSION AND NUCLEOTIDE-SEQUENCE OF A THERMOSTABLE DNA LIGASE-ENCODING GENE

被引：74

作者：

BARANY, F ^{[1
]}

GELFAND, DH ^{[1
]}

机构：

[1] CETUS CORP, PCR DIV, EMERYVILLE, CA 94608 USA

来源：

GENE | 1991年 / 109卷 / 01期

关键词：

LIGASE CHAIN REACTION; MRRB; PHOA PROMOTER; PROTEIN-DNA INTERACTIONS; RECOMBINANT DNA;

D O I：

10.1016/0378-1119(91)90582-V

中图分类号：

Q3 [遗传学];

学科分类号：

071007 ; 090102 ;

摘要：

Thermostable DNA ligase has been harnessed for the detection of single-base genetic diseases using the ligase chain reaction [Barany, Proc. Natl. Acad. Sci. USA 88 (1991) 189-193]. The Thermus thermophilus (Tth) DNA ligase-encoding gene (ligT) was cloned in Escherichia coli by genetic complementation of a ligts 7 defect in an E. coli host. Nucleotide sequence analysis of the gene revealed a single chain of 676 amino acid residues with 47% identity to the E. coli ligase. Under phoA promoter control, Tth ligase was overproduced to greater than 10% of E. coli cellular proteins. Adenylated and deadenylated forms of the purified enzyme were distinguished by apparent molecular weights of 81 kDa and 78 kDa, respectively, after separation via sodium dodecyl sulfate-polyacrylamide-gel electrophoresis.

引用

页码：1 / 11

页数：11

共 73 条

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