中国农业科学 ›› 2011, Vol. 44 ›› Issue (19): 4096-4101.doi: 10.3864/j.issn.0578-1752.2011.19.021

• 兽医 • 上一篇    下一篇

斯氏艾美耳球虫MIC-5与兔IL-15基因在真核细胞中的共表达

孟庆玲, 乔军, 才学鹏, 田广孚, 闫鸿斌, 骆学农   

  1. 1.中国农业科学院兰州兽医研究所/家畜疫病病原生物学国家重点实验室
    2.石河子大学动物科技学院
  • 收稿日期:2010-03-29 出版日期:2011-10-01 发布日期:2011-08-23
  • 通讯作者: 通信作者才学鹏,E-mail:caixp@public.lz.gs.cn
  • 作者简介:孟庆玲,Tel:13779353695;E-mail:xjmqlqj@sina.com
  • 基金资助:

    科技部家畜疫病病原生物学国家重点实验室项目(2008-001)、石河子大学高层次人才专项基金(2009-011)

Co-expression of MIC-5 Gene of Eimeria stiedai and Rabbit IL-15 in Eucaryotic Cell Line

 MENG  Qing-Ling, QIAO  Jun, CAI  Xue-Peng, TIAN  Guang-Fu, YAN  Hong-Bin, LUO  Xue-Nong   

  1. 1.中国农业科学院兰州兽医研究所/家畜疫病病原生物学国家重点实验室
    2.石河子大学动物科技学院
  • Received:2010-03-29 Online:2011-10-01 Published:2011-08-23

摘要: 【目的】构建能在真核细胞中共表达斯氏艾美耳球虫微线蛋白5(MIC-5)和兔IL-15重组载体,为进一步研究抗斯氏艾美耳球虫的免疫调节性DNA疫苗奠定基础。【方法】将斯氏艾美耳球虫MIC-5和兔IL-15基因,克隆到真核双表达载体pBudce4.1中,应用脂质体介导法将构建的重组质粒pB-MIC-5-IL-15转染BHK-21细胞。在转染后,用RT-PCR法和间接免疫荧光试验(IFA)分别检测MIC-5和IL-15基因在BHK-21细胞中的转录和表达情况。【结果】在重组质粒pB-MIC-5-IL-15转染BHK-21细胞24 h后即可检测到MIC-5、IL-15基因进行了转录;在转染36、48、72h后,MIC-5和IL-15基因可同时在BHK-21细胞中表达。【结论】成功构建了含有斯氏艾美耳球虫MIC-5和兔IL-15基因的重组载体,实现了两种基因在真核细胞中的共表达,为抗斯氏艾美耳球虫的免疫调节性DNA疫苗研制奠定了基础。

关键词:

Abstract: 【Objective】 The aim of this study is to construct a recombinant co-expression vector expressing Eimeria stiedai microneme protein 5 (MIC-5) and rabbit IL-15 in eukaryotic cells, which may lay a foundation for further study immune regulatory DNA vaccine against infection of Eimeria stiedai. 【Method】 E. stiedai MIC-5 gene and rabbit's IL-15 gene were respectively cloned into plasmid pBudce4.1 to construct recombinant vector pB-MIC-5-IL-15 for co-expression, and then the recombinant plasmid pB-MIC-5-IL-15 was transfected into BHK-21 cell line for expression. The RT-PCR and indirect fluorescent immunoassay were used to detect the transcription and expression of interesting genes in BHK-21 cell line, respectively. 【Result】 The results showed that E. stiedai MIC-5 gene and rabbit's IL-15 gene  were successfully transcripted. And interesting genes of E.stiedai MIC-5 and rabbit's IL-15 were successfully expressed at 36, 48 and 72 hour post-transfection, respectively. 【Conclusion】 The recombinant co-expression vector was successfully constructed, which can express E. stiedai MIC-5 and rabbit IL-15 gene in eukaryotic cells.

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