Feutrier J, Kay W W, Trust T J
Department of Biochemistry and Microbiology, University of Victoria, British Columbia, Canada.
J Bacteriol. 1988 Sep;170(9):4216-22. doi: 10.1128/jb.170.9.4216-4222.1988.
A gene bank of DNA from a human isolate of Salmonella enteritidis was constructed in the cosmid pHC79 in Escherichia coli HB101. Five clones containing 35- to 45-kilobase inserts of S. enteritidis DNA reacted in colony immunoblot assays with a polyclonal antiserum prepared against purified S. enteritidis fimbriae. Electron microscopy showed that none of the five fimbrin-producing clones produced fimbriae, yet radioimmunoprecipitation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis located the 14,400-molecular-weight S. enteritidis in the outer membrane fraction of three of the clones and in the periplasmic fraction of all five clones. By using an oligonucleotide probe homologous to the 5' region of the fimbrin structural gene, the fimbrin gene was located on a 5.3-kilobase HindIII fragment. In vitro transcription-translation analysis verified that this HindIII fragment subcloned into plasmid pTZ18R produced unprocessed S. enteritidis fimbrin of molecular weight 16,400. Dot blot hybridization against a selection of strains of the family Enterobacteriaceae indicated a limited distribution of the S. enteritidis fimbrin gene.
在大肠杆菌HB101中，利用黏粒pHC79构建了肠炎沙门氏菌人分离株的DNA基因文库。五个含有35至45千碱基肠炎沙门氏菌DNA插入片段的克隆，在菌落免疫印迹试验中与针对纯化的肠炎沙门氏菌菌毛制备的多克隆抗血清发生反应。电子显微镜显示，这五个产生菌毛蛋白的克隆中没有一个产生菌毛，但放射免疫沉淀和十二烷基硫酸钠-聚丙烯酰胺凝胶电泳表明，分子量为14400的肠炎沙门氏菌菌毛蛋白存在于三个克隆的外膜组分中以及所有五个克隆的周质组分中。通过使用与菌毛蛋白结构基因5'区域同源的寡核苷酸探针，菌毛蛋白基因位于一个5.3千碱基的HindIII片段上。体外转录-翻译分析证实，亚克隆到质粒pTZ18R中的这个HindIII片段产生了分子量为16400的未加工肠炎沙门氏菌菌毛蛋白。针对肠杆菌科的一系列菌株进行的点杂交表明，肠炎沙门氏菌菌毛蛋白基因的分布有限。
