Cloning,Expression and Immunogenicity Analysis of OmpU of Vibrio harveyi ZJ 2008
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    Abstract:

    According to OmpU gene sequence published in GenBank,primers were designed and the DNA fragment of about 1 000 bp was amplified by PCR from genomic DNA of Vibrio Harveyi ZJ2008,isolated from disease Pseudosciaena crocea.The gene was cloned into pMD18-T vector and sequenced,then submitted to GenBank with an accession number FJ919231.BlastN result showed that the sequence shared identity of 99.0% with OmpU of Vibrio harveyi.The sequence was subcloned into pET-30a resulted with the recombinant plasmid pET30aOmpU which was further transferred into host bacteria E.coli BL21(DE3) with IPTG induction for expression.SDSPAGE profiles showed the expected MW of the recombinant OmpU was around 41 ku.The protein was expressed in the form of insolvable fusion bodies and was preliminary purified by urea method.Forty pieces of large yellow croaker were individually immunized with 50 μg of the recombinant protein by intraperitoneal injection.Four weeks after vaccination,ten pieces were artificially challenged by intraperitoneal injection with live bacteria and the mortality was recorded in the following 14 d.Relative percentage of the survival rate of the immunized group reached 85%.Specific antibody level was detected by indirect ELISA after 4-8 weeks post vaccination.Resulting an increase of antibody titers to the immunized recombinant protein.The present study demonstrated strong immunogenicity of the recombinant OmpU.OmpU of V.harveyi should be given enough attention for their potential application in vaccine development.

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黄辉,毛芝娟,陈吉刚. Cloning, Expression and Immunogenicity Analysis of OmpU of Vibrio harveyi ZJ 2008[J]. Jorunal of Huazhong Agricultural University,2010,29(3):346-350.

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  • Received:January 05,2010
  • Revised:March 01,2010
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