›› 2012, Vol. 39 ›› Issue (11): 43-46.

• 生物技术 • 上一篇    下一篇

奶牛载脂蛋白B100基因克隆及原核表达

张加力1,2, 王力2, 杨文涛1, 丁红研1, 张良1, 陈灰1, 刘兆喜1, 赵晨旭1, 王哲1, 李小兵1   

  1. 1. 吉林大学畜牧兽医学院,吉林长春 130062;
    2. 吉林农业大学动物科学技术学院,吉林长春 130118
  • 收稿日期:2012-04-08 出版日期:2012-11-20 发布日期:2012-11-22
  • 通讯作者: 李小兵。E-mail:xbli@jlu.edu.cn E-mail:xbli@jlu.edu.cn
  • 作者简介:张加力(1972-),男,吉林人,副教授,主要从事动物营养代谢障碍性疾病研究。
  • 基金资助:
    国家自然科学基金项目(30972224)。

Cloning and Prokaryotic Expression of Apolipoprotein B100 Gene of Dairy Cows

ZHANG Jia-li1,2, WANG Li2, YANG Wen-tao1, DING Hong-yan1, ZHANG Liang1, CHEN Hui1, LIU Zhao-xi1, ZHAO Chen-xu1, WANG Zhe1, LI Xiao-bing1   

  1. 1. College of Animal Science and Veterinary Medicine,Jilin University,Changchun 130062,China;
    2. Department of Animal Science and Technology,Jilin Agricultural University,Changchun 130118,China
  • Received:2012-04-08 Online:2012-11-20 Published:2012-11-22

摘要: 本研究根据GenBank已收录奶牛ApoB100基因序列,设计特异性引物获得目的基因。经pGM-T载体克隆,对重组质粒进行BamHⅠ和EcoRⅠ双酶切鉴定并测序,序列同源性达100%;将目的基因连接到pET-28a表达载体中,提取质粒,转化到Rosetta(DE3)中,筛选的阳性克隆经IPTG诱导。SDS-PAGE初步分析表明,成功获得分子质量为35 ku的蛋白质;Western blotting结果呈阳性,表明通过本试验成功获得了目的蛋白。

关键词: ApoB100; 基因克隆; 原核表达

Abstract: Specific primers were designed according to ApoB100 gene sequence reported in GenBank in this study. Cloning was carried out by the pGM-T vector, recombined plasmid DNA was cut by BamHⅠand EcoRⅠenzymes and then sequencing. The recombinant expression vector pET-28a-ApoB100 was constructed with target gene and pET-28a vector, and transformed into Rosetta(DE3),then induced by IPTG. The expression product was identified by SDS-PAGE and Western blotting. The results showed that the homology of the cloned ApoB100 gene was 100% to that reported in GenBank. SDS-PAGE analysis showed that the molecular weight of recombinant protein pET-28a-ApoB100 was 35 ku. Western blotting positive results showed that the experiment successfully obtained the target protein.

Key words: ApoB100; gene cloning; prokaryotic expression

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