Stanley K K, Luzio J P
EMBO J. 1984 Jun;3(6):1429-34. doi: 10.1002/j.1460-2075.1984.tb01988.x.
Construction of a family of bacterial expression vectors, pEX1-3, is described. These vectors are derived from a cro-lacZ gene fusion plasmid which expresses large quantities of fusion protein under the control of the PR promoter of bacteriophage lambda. A polylinker has been engineered into the 3' end of the lacZ gene in all three translational reading frames, and stop signals for transcription and translation inserted, so that any open reading frame DNA may be expressed as a hybrid beta-galactosidase protein. cDNA fragments cloned in these vectors can be detected with an efficiency of greater than 1 in 3, thus enabling the detection of rare cDNA molecules. In addition, the low solubility of hybrid proteins leads to a rapid isolation procedure allowing antibodies of pre-determined specificity to be made against expressed regions of cloned DNA. We describe the cloning of albumin and complement C9 genes from a human cDNA library using polyclonal and monoclonal antibodies.
本文描述了一系列细菌表达载体pEX1 - 3的构建。这些载体源自一种cro - lacZ基因融合质粒，该质粒在噬菌体λ的PR启动子控制下可表达大量融合蛋白。在lacZ基因的3'端的所有三个翻译阅读框中都设计了一个多克隆位点，并插入了转录和翻译的终止信号，这样任何开放阅读框DNA都可以作为杂合β - 半乳糖苷酶蛋白来表达。克隆到这些载体中的cDNA片段的检测效率大于三分之一，从而能够检测到罕见的cDNA分子。此外，杂合蛋白的低溶解性导致了一种快速的分离程序，使得能够针对克隆DNA的表达区域制备具有预定特异性的抗体。我们描述了使用多克隆抗体和单克隆抗体从人cDNA文库中克隆白蛋白和补体C9基因的过程。
