摘要
通过RT-PCR扩增禽呼肠孤病毒σC基因,经EcoRⅠ和SalⅠ双酶切处理后与pET-30a原核表达载体连接,获得重组质粒pET-30a-σC。将重组阳性质粒转化至BL21(DE3)感受态细胞,经IPTG诱导表达后通过Western blot分析表明重组目的蛋白获得表达,分子量约为42ku,主要以包涵体形式存在。重组蛋白经纯化后免疫新西兰白兔,三次免疫获得抗σC蛋白的高效免疫血清。经ELISA检测,该血清抗体效价为1∶105,并且能够与重组杆状病毒表达的σC蛋白发生特异性反应。禽呼肠孤病毒σC蛋白的表达与兔抗血清的制备为进一步研究该蛋白功能奠定了基础。
σC gene of avian reovirus was amplified by RT-PCR and cloned into prokaryotic expression vector pET-30a based on restriction enzyme analysis. The recombinant expression vector pET-30a-σC was transformed into the competent BL21 （DE3）. The fusion protein was expressed after induced by IPTG and presented mainly in inclusion body. Western blot analysis showed that the fusion protein （42 ku）was expressed correctly. The specific antiserum against σC was produced in rabbit immunized three times by the purified fusion protein σC. The antibody titer was 1∶105 in ELISA assay,and the antibody had specific reaction with the protein σC expressed by recombinant Baculovirus. The production of σC protein and antiserum will be helpful for function research of σC protein.
出处
《中国家禽》
北大核心
2011年第12期18-21,共4页
China Poultry
基金
现代农业产业技术体系建设专项资金(nycytx-42-G3-01)
