| 猪Fc受体γ亚单位基因克隆与原核表达 |
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| 引用本文： | 张志远,张继希,徐红运,刘肖萍,卢晓艳,段二珍,夏平安,崔保安.猪Fc受体γ亚单位基因克隆与原核表达[J].华北农学报,2012(1):35-38. |
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| 作者姓名： | 张志远 张继希 徐红运 刘肖萍 卢晓艳 段二珍 夏平安 崔保安 |
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| 作者单位： | 河南农业大学牧医工程学院 |
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| 基金项目： | 国家自然科学基金项目(31172346) |
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| 摘 要： | 应用RT-PCR技术从90日龄健康仔猪肺巨噬细胞中克隆出猪Fcγ亚单位的cDNA序列,并将其亚克隆到原核表达载体pET-32a中,构建重组表达质粒pET-FcRγ,成功表达了分子量约为29 kDa的γ亚单位重组蛋白。将重组蛋白FcRγ-His免疫小鼠,制备获得鼠抗FcRγ-His重组蛋白多克隆抗体,将得到的重组蛋白多克隆抗体采用间接ELISA和Western-blotting试验方法检测,ELISA测定多克隆抗体的效价为1∶8 000。Western-blotting结果表明,制备的鼠抗FcRγ-His重组蛋白多克隆抗体可以与重组γ亚单位蛋白进行特异性结合,从而证明重组蛋白具有较好的免疫原性。成功克隆出猪Fc受体γ亚单位基因并表达,为进一步研究其结构与功能奠定了基础。
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| 关 键 词： | 猪Fc受体γ单位 克隆 原核表达 多克隆抗体 |
Gene Cloning of Porcine Fc Receptor Gamma Subunit and Its Prokaryotic Expression |
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| ZHANG Zhi-yuan,ZHANG Ji-xi,XU Hong-yun,LIU Xiao-ping, LU Xiao-yan,DUAN Er-zhen,XIA Ping-an,CUI Bao-an.Gene Cloning of Porcine Fc Receptor Gamma Subunit and Its Prokaryotic Expression[J].Acta Agriculturae Boreali-Sinica,2012(1):35-38. |
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| Authors: | ZHANG Zhi-yuan ZHANG Ji-xi XU Hong-yun LIU Xiao-ping LU Xiao-yan DUAN Er-zhen XIA Ping-an CUI Bao-an |
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| Institution: | (College of Animal Science and Veterinary Medicine,Henan Agricultural University,Zhengzhou 450002,China) |
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| Abstract: | mRNA encoding porcine FcR gamma subunit obtained from PAMs of 90-day piglets was transcribed into cDNA through RT-PCR.Recombinant plasmid pET-FcRγ was constructed successfully by inserting the cDNA sequence into pET-32a,and expressed as soluble protein with a molecular weight of about 29 kDa.Following purification,recombinant protein FcRγ-His was used to immune mouse,and polyclonal antibody against it was obtained.Western-blotting indicated that such polyclonal antibody could react specifically with FcRγ-His.Indirect ELISA shows that the titer of the polyclonal antibody is 1 ∶ 8000,revealing the good immunogenicity of FcRγ-His.All these results provide the foundation for further studying the structure and function of FcR gamma subunit. |
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| Keywords: | Porcine FcR gamma subunit Gene clone Prokaryotic expression Polyclonal antibody |
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