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[普氏立克次氏体三个编码外膜蛋白的基因在大肠杆菌中的克隆与表达]

[Cloning and expression in Escherichia coli of three Rickettsia prowazekii genes, coding outer membrane proteins].

作者信息

Emel'ianov V V, Balaeva N M

出版信息

Mol Biol (Mosk). 1992 Jul-Aug;26(4):847-58.

PMID:1279381
Abstract

Rickettsia prowazekii (virulent Breinl strain) random genomic DNA fragments were cloned in the lambda gt11 expression vector by using non-palindromic adaptors. Several immunoreactive clones were selected after screening 20,000 individual recombinant plaques with human convalescent serum. Some recombinants synthesized the complete 60 K protein, and others synthesized beta-galactosidase fusion polypeptides containing epitopes of 134 K protein of the R. prowazekii outer membrane. The amplified genomic library was screened with monospecific antibodies directed against abundant 31 K and 29.5 K outer membrane proteins. Several recombinant clones expressing full or part of 29.5 K polypeptide, and none expressing 31 K polypeptide were revealed. The serum of a patient convalescing from epidemic typhus did not react in western blot with recombinant 29.5 K protein.

摘要

用非回文衔接头将普氏立克次体(强毒株Breinl株)随机基因组DNA片段克隆于λgt11表达载体。用人恢复期血清筛选20000个单个重组噬菌斑后,挑选出几个免疫反应性克隆。一些重组体合成完整的60K蛋白,另一些合成含普氏立克次体外膜134K蛋白表位的β-半乳糖苷酶融合多肽。用针对丰富的31K和29.5K外膜蛋白的单特异性抗体筛选扩增的基因组文库。发现了几个表达完整或部分29.5K多肽的重组克隆,未发现表达31K多肽的克隆。一名斑疹伤寒恢复期患者的血清在蛋白质印迹中与重组29.5K蛋白无反应。

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