鸡γ-干扰素成熟肽基因的克隆、表达与抗血清制备  

Cloning and expression of mature peptide gene of chicken IFN-γ and preparation of antiserum

作　　者：王永娟[1] 朱善元[1] 左伟勇[1] 王安平[1] 吴双[1] 洪伟鸣[1] 黄文强[1] 韩小英[1] 

机构地区：[1]江苏畜牧兽医职业技术学院,江苏省兽用生物制药高技术研究重点实验室,江苏泰州225300

出　　处：《江苏农业学报》2012年第6期1373-1377,共5页Jiangsu Journal of Agricultural Sciences

基　　金：2012年江苏畜牧兽医职业技术学院重点支持项目(ZD1204);2010年江苏省教育厅科研成果产业化推进项目(JHZD09-71);江苏畜牧兽医职业技术学院博士启动基金项目(ZD200908)

摘　　要：为了制备安全、高效的重组鸡γ-干扰素及抗血清,以PCR方法从活化的鸡外周血淋巴细胞中扩增鸡γ-干扰素的成熟肽基因序列,并构建原核表达质粒pET32a-γ和pGEX6p-γ;SDS-PAGE法检测2个载体特异性表达成熟肽的情况;将纯化的pET32a-γ诱导表达产物免疫小鼠,收获小鼠血清,并以pGEX6p-γ诱导表达产物检测其中特异性抗体滴度。结果显示,扩增的鸡γ-干扰素成熟肽基因能成功表达,原核表达产物免疫小鼠可产生特异性抗体,抗体效价为1∶16 000。表明所用的表达系统可以制备安全、高效的重组干扰素,能够为定量检测机体内产生的γ-干扰素进而了解机体针对病原的免疫状态提供较为可靠的技术手段,为其他一些尚无商品化抗体的生物分子鉴定提供参考方法。To prepare safe and efficient recombinant chicken IFN-γ and its antiserum,the gene sequence of mature peptide of chicken IFN-γ was amplified from activated chicken peripheral blood lymphocytes by PCR.pET32a-γ and pGEX6p-γ were constructed for mature IFN-γ expression and dectected by SDS-PAGE.Purified pET32a-γ-induced protein was injected into mice,and peptide-specific antibody was detected by pGEX6p-γ-induced protein.Results showed that the gene of mature peptide of chicken IFN-γ was successfully expressed via prokaryotic expression vector,and immunized mice successfully produced specific antibody,with the antibody titer of 1∶16 000.The study provided a more reliable technique for quantitative detection of IFN-γ and other biomolecules with no commercial antibodies.

关 键 词：鸡 Γ-干扰素 原核表达 抗体 

分 类 号：S831.49[农业科学—畜牧学]