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首页> 外文期刊>Journal of bacteriology >Cloning and characterization of the Salmonella typhimurium metE gene.
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Cloning and characterization of the Salmonella typhimurium metE gene.

机译：鼠伤寒沙门氏菌metE基因的克隆与鉴定。

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摘要

The metE gene of Salmonella typhimurium was cloned into plasmid pACYC184 by using a lambda gt7 -metE transducing phage as a source of metE DNA. The recombinant plasmid, designated pGS41 , carries a 12.8-kilobase-pair EcoRI insert fragment. The metE gene was subcloned from pGS41 into plasmid pBR322 on a 4.2-kilobase-pair EcoRI-HindIII fragment (plasmid pGS47 ) and a 4.5-kilobase-pair PstI fragment (plasmid pGS69 ). The location of metE in these plasmids was determined by transposon Tn5 insertional inactivation experiments. A metE-encoded polypeptide of 92,500 Mr was detected in a minicell system by using metE+ plasmids as templates. Truncated polypeptides replaced the 92,500-Mr polypeptide when plasmid derivatives containing Tn5 insertions that inactivate metE were used in the system. A comparison of the site of each Tn5 insertion and the size of the polypeptide made in the minicell system allowed us to determine the direction of transcription and translation. The location of the metE promoter was determined by using an in vitro transcription system and by RNA polymerase protection of restriction endonuclease sites.
机译：通过λgt7-metE转导噬菌体作为metE DNA的来源，将鼠伤寒沙门氏菌的metE基因克隆到质粒pACYC184中。命名为pGS41的重组质粒带有一个12.8碱基对的EcoRI插入片段。将metE基因从pGS41亚克隆到4.2碱基对的EcoRI-HindIII片段（质粒pGS47）和4.5碱基对的PstI片段（质粒pGS69）上的质粒pBR322中。通过转座子Tn5插入失活实验确定metE在这些质粒中的位置。通过使用metE +质粒作为模板，在小细胞系统中检测到92,500 Mr的metE编码多肽。当在系统中使用含有灭活metE的Tn5插入的质粒衍生物时，截短的多肽取代了92,500-Mr多肽。通过比较每个Tn5插入的位点和在小细胞系统中制备的多肽的大小，我们可以确定转录和翻译的方向。 metE启动子的位置是通过使用体外转录系统和限制性内切核酸酶位点的RNA聚合酶保护来确定的。

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