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    吕平, 彭万胜, 董淮富. IL-35重组卡介苗的构建及鉴定[J]. 蚌埠医科大学学报, 2020, 45(7): 849-853, 858. DOI: 10.13898/j.cnki.issn.1000-2020.07.003
    引用本文: 吕平, 彭万胜, 董淮富. IL-35重组卡介苗的构建及鉴定[J]. 蚌埠医科大学学报, 2020, 45(7): 849-853, 858. DOI: 10.13898/j.cnki.issn.1000-2020.07.003
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    LÜ Ping, PENG Wan-sheng, DONG Huai-fu. Construction and identification of the IL-35-BCG recombinant vaccine[J]. Journal of Bengbu Medical University, 2020, 45(7): 849-853, 858. DOI: 10.13898/j.cnki.issn.1000-2020.07.003
    Citation: LÜ Ping, PENG Wan-sheng, DONG Huai-fu. Construction and identification of the IL-35-BCG recombinant vaccine[J]. Journal of Bengbu Medical University, 2020, 45(7): 849-853, 858. DOI: 10.13898/j.cnki.issn.1000-2020.07.003
    shu

    IL-35重组卡介苗的构建及鉴定

    Construction and identification of the IL-35-BCG recombinant vaccine

      摘要
    • 摘要:
      目的构建分泌性表达白细胞介素（IL）-35真核载体pZM03（pBudCE4.1/GLS/IL35/OriM）-重组卡介苗（BCG）。
      方法在化学合成的IL-35基因片段中插入pBudCE4.1/GLS/OriM，将合成后序列插入到IL-12真核共表达载体pBudCE4.1/GLS/IL12/OriM的质粒酶切回收产物pBudCE4.1/GLS/OriM中，构建表达IL-35的穿梭/共表达质粒重组质粒pZM03（pBudCE4.1/GLS/IL35/OriM），再以重组质粒电转化到BCG，构建表达IL-35和BCG的重组pBudCE4.1/GLS/IL35/OriM-BCG。
      结果重组质粒pZM03（pBudCE4.1/GLS/IL35/OriM）经PCR扩增、测序鉴定，以及在CHO 293T细胞中表达产物的Western blotting检测，从转录和翻译水平对目标基因产物进行检测验证，pBudCE4.1/GLS/IL35/OriM-BCG在细胞水平经抗酸染色验证，证实基因IL-35正确插入载体pBudCE4.1/GLS/OriM中，且pBudCE4.1/GLS/IL35/OriM-BCG构建成功。
      结论重组质粒pBudCE4.1/GLS/IL35/OriM-BCG可正确表达IL-35，为进一步研究IL-35功能及其应用价值奠定了良好的基础。

       

      Abstract:
      ObjectiveTo construct the recombinant eukaryotic expression vector pZM03 (pBudCE4.1/GLS/IL35/OriM) containing secretory protein of interleukin 35 (IL-35)-bacillus calmette guerin (BCG)recombinant vaccine.
      MethodsThe chemically synthesized IL-35 gene fragment was cloned into the pBudCE4.1/GLS/OriM, the pBudCE4.1/GLS/IL35/OriM was obtained from the pBudCE4.1/GLS/IL12/OriM, and both of above was used to construct the co-expression vector pZM03 (pBudCE4.1/GLS/IL35/OriM).The recombinant plasmid pZM03 was electrotransformated into the BCG to construct the recombinant co-expression of IL-35 and BCG plasmid pBudCE4.1/GLS/IL35/OriM-BCG.
      ResultsThe results of the detection of gene, transcription, translation and cellular levels, which included the PCR, sequencing, Western blotting and acid-fast staining methods, showed that the pBudCE4.1/GLS/IL35/OriM-BCG was successfully constructed.
      ConclusionThe recombinant plasmid pBudCE4.1/GLS/IL35/OriM-BCG can correctly express IL-35, which lays a good foundation for further research on the function and application value of IL-35.

       

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