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《Journal of First Military Medical University》 2001-02
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Cloning of thermostable direct hemolysin gene of Vibrio parahaemolyticus

WU Zhen-long, NIE Jun, BAO Hui-ning, XU Bing, WANG Hong (Department of Epideminlogy, First Military Medical University, Guangzhou 510515,China ;Tongling People's Hospital of Anhui Province,Tongling 244000, China; Institute of Biotechnology, Academy o f Mi  
Objective To clone thermostable direct hemolysin (TDH)gene (tdh gene) of Vibrio parahaemolyticus. Methods PCR technique was adopted to amplify the target fragments of tdh gene from the genomic DNA of Vp14-90. Digestion with the restriction endonucleases EcoRI and BamHI was performed and the digested fragments were ligated with plasmid pUC19 followed by digestion in the same manner. The recombinant plasmid was subsequently transformed into E.coli JM109, and the positive clone was screened and identified by PCR and restriction analysis. Sequencing of the recombinant plasmid was con- ducted. Rasults DNA agarose gel electrophoresis showed that the PCR product of Vp14-90 was about 600 bp, confirmed by PCR and double restriction analysis of the positive clone. The homogneity between the positive clone and the reported tdh gene in Genebank was 99.65%. Conxlusion The positive recombinant clone were successfully cloned. which has laid preliminary foundation for studying the pathogenesity of Vibrio parahaemolyticus and also for preparing specific gene probe for the bacterial detection.
【CateGory Index】: R394.8
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