畜牧与饲料科学 ›› 2014, Vol. 35 ›› Issue (12): 1-1.doi: 10.12160/j.issn.1672-5190.2014.12.003

• 基础科学 • 上一篇    下一篇

猪干扰素α-1基因的原核表达与多克隆抗体制备

刘娜 李春秋 高晶 耿雨菲 王欣宇 苏明俊 魏姗 王志慧 王建发 武瑞 孙东波   

  1. 黑龙江八一农垦大学动物科技学院,黑龙江大庆163319
  • 出版日期:2014-12-20 发布日期:2014-12-20
  • 通讯作者: 刘娜
  • 作者简介:刘娜(1988-),女,硕士研究生,主要研究方向为兽医临床微生物。 通讯作者:孙东波(1978-),男,副教授,博士,硕士生导师,主要研究方向为兽医临床微生物与奶牛肢蹄病。
  • 基金资助:
    黑龙江省高校新世纪优秀人才项目(1252-NCET-016)。

Prokaryotic Expression of Porcine Interferonα-1 and Preparation of Polyclonal Antibodies

LIU Na,LI Chun-qiu,GAO Jing,GENG Yu-fei,WANG Xin-yu,SU Ming-jun,WEI Shan,WANG Zhi-hui,WANG Jian-fa,WU Rui,SUN Dong-bo(College of Animal Science and Technology, Heilongjiang Bayi Agricultural University, Daqing 163319, China)   

  • Online:2014-12-20 Published:2014-12-20

摘要: [目的]利用大肠杆菌原核表达系统对猪干扰素α-1(PoIFNα-1)基因进行表达,制备PoIFNα-1重组蛋白多克隆抗体。[方法]根据GenBank发表的PoIFNα-1核苷酸序列,合成密码子优化的PoIFNα-1基因,构建PoIFNα-1基因的pET-28a原核表达载体,转化大肠杆菌BL21(DE3)宿主菌后,利用IPTG进行诱导表达。表达的PoIFNα-1重组蛋白纯化后,免疫昆明系小鼠,制备多克隆抗体。采用Western blot技术检测PoIFNα-1重组蛋白的多克隆抗体与天然PoIFNα-1的反应性。[结果]合成的PoIFNα-1基因在大肠杆菌BL21(DE3)宿主菌中成功表达,针对PoIFNα-1重组蛋白的多克隆抗体能够识别天然的猪干扰素α-1。[结论]合成的PoIFNα-1基因在大肠杆菌中获得了成功表达,为后续研究奠定了基础。

Abstract: Objective] The porcine interferon alpha 1 gene (PoIFNα-1) was expressed by using Escherichia coli prokaryotic expression system, and polyclonal antibodies against the recombinant protein PoIFNα-1 were prepared. [Method] The codon-optimized PoIFNα-1 gene was synthesized according to the nucleotide sequence published by GenBank, and then the synthesized PoIFNα-1 gene was cloned into the vector pET-28a. After transformation of the recombinant plasmid into E. coli BL21 (DE3) host bacteria, the recombinant bacterium was induced by using IPTG. After purification of the recombinant protein PoIFNα-1 expressed in E. coli BL21 (DE3), the polyclonal antibodies against the recombinant protein PoIFNα-1 was prepared in Kunming mice, and then reactivity of the prepared polyclonal antibodies with native porcine interferon alpha 1 was determined by Western blot. [Result] The synthesized PoIFNα-1 gene was successfully expressed in E. coli BL21 (DE3), and the polyclonal antibody against recombinant protein PoIFNα-1 could recognize the native porcine interferon-α-1. [Conclusion] The synthesized PoIFNα-1 gene was successfully expressed in E. coli, which provided some basis for following further research of antivirus.

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