INVIVO OVEREXPRESSION AND PURIFICATION OF ESCHERICHIA-COLI TRANSFER RNA(SER)

被引：16

作者：

BOREL, F ^{[1
]}

HARTLEIN, M ^{[1
]}

LEBERMAN, R ^{[1
]}

机构：

[1] EUROPEAN MOLEC BIOL LAB,GRENOBLE OUTSTN,156X,F-38042 GRENOBLE,FRANCE

来源：

FEBS LETTERS | 1993年 / 324卷 / 02期

关键词：

SERINE; TRANSFER RNA; AMINOACYL-TRANSFER RNA SYNTHETASE; OVEREXPRESSION;

D O I：

10.1016/0014-5793(93)81385-D

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

DNA fragments corresponding to the sequences of Escherichia coli tRNA2ser and amber suppressor tRNA(ser), were synthesized from overlapping oligonucleotides. These were interposed between a strong promotor and a synthetic transcriptional terminator to ensure the production of a transcript of the correct size. The genes of promotor, fragment and terminator were cloned into a conditional runaway replication plasmid. At temperatures below 37-degrees-C this vector has a low copy number but, following a temperature shift to 42-degrees-C, the copy number is no longer regulated. Using these constructs an overexpression of tRNA(ser) of about 20 times the level of the wild-type pool could be obtained (corresponding e.g. to 200 times the expression tRNA2ser). From these systems 10 mg quantities of tRNA(ser)s could be isolated with a serine acceptance of 1,100 pmol/A280) unit.

引用

页码：162 / 166

页数：5

共 20 条

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