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pJC20和pJC40——用于在大肠杆菌中T7 RNA聚合酶依赖性表达重组基因的两种高拷贝数载体。

pJC20 and pJC40--two high-copy-number vectors for T7 RNA polymerase-dependent expression of recombinant genes in Escherichia coli.

作者信息

Clos J, Brandau S

机构信息

Bernhard Nocht Institute for Tropical Medicine, Hamburg, Germany.

出版信息

Protein Expr Purif. 1994 Apr;5(2):133-7. doi: 10.1006/prep.1994.1020.

Abstract

We report the construction of two plasmid vectors, pJC20 and pJC40, for the expression of recombinant genes in Escherichia coli under the control of T7 RNA polymerase. Their small sizes of ca. 2.4 kb ease the subcloning of large inserts and the high copy numbers obtained result in satisfactory yields in all plasmid preparations. A multiple-cloning site offers sites for directional cloning and nested deletions. In addition, pJC40 encodes a cleavable amino-terminal histidine tail of 10 residues which is added to the gene product, thus allowing purification by metal chelate chromatography. Observed expression yields are in the range of 10% of total bacterial protein for all genes tested in our laboratory.

摘要

我们报道了两种质粒载体pJC20和pJC40的构建,它们用于在T7 RNA聚合酶控制下在大肠杆菌中表达重组基因。它们约2.4 kb的小尺寸便于大片段的亚克隆,并且获得的高拷贝数在所有质粒制备中都能产生令人满意的产量。一个多克隆位点提供了用于定向克隆和嵌套缺失的位点。此外,pJC40编码一个可切割的10个残基的氨基末端组氨酸尾巴,该尾巴被添加到基因产物中,从而允许通过金属螯合层析进行纯化。在我们实验室测试的所有基因中,观察到的表达产量在总细菌蛋白的10%范围内。

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