肉鸡HSP70基因克隆、原核表达及单克隆抗体制备
lone and expression of broiler HSP70 gene in E. coli and preparation of HSP70 monoclonal antibody
利用RT-PCR方法扩增了肉鸡HSP70 mRNA，将其扩增产物克隆到PGEM-T Easy 载体和原核表达载体pET-28a （+）上，进一步转化至大肠杆菌BL-21中，用IPTG诱导表达重组肉鸡HSP70。以纯化的重组肉鸡HSP70制备多克隆抗体，免疫印迹分析说明重组肉鸡HSP70具有很好的免疫原性。以纯化的重组肉鸡HSP70免疫BALB/c小鼠，通过淋巴细胞杂交瘤技术，获得2株分泌肉鸡HSP70单克隆抗体的杂交瘤细胞株，进一步通过免疫印迹分析筛选出1株高特异性的肉鸡HSP70单克隆抗体，命名为BH70-1。免疫组织化学结果显示，单克隆抗体BH70-1具有很好的组织亲和力，是一种理想的抗体。
he mRNA of broiler HSP70 gene was amplified by RT-PCR and the production was cloned into the PGEM-T Easy vector and the expression vector pET-28a（+）. The recombinant expression vector was transformed into E. coli BL-21 and induced by IPTG to express. The recombinant broiler HSP70 was purified to prepare the polyclonal antibody and the immunel activity of recombinant broiler HSP70 was proved by western-blot. The recombinant broiler HSP70 was used to immunize BALB/ c mice and two hyridoma cell lines secreting monoclonal antibody of broiler HSP70 were established by hybridoma technique. One of the high-specific monoclonal antibody for broiler HSP70 was selected by western-blot and named as BH70-1. The immunohistochemistry results showed that the monoclonal antibody BH70-1 has high affinity with broiler tissues. The results suggest that the prepared monoclonal antibody BH70-1 is effective.
王志亮、吴晓东、鲍恩东、刘雨田、孙培明、赵永刚
生物科学研究方法、生物科学研究技术分子生物学生物工程学
热休克蛋白70原核表达单克隆抗体免疫印迹免疫组织化学肉鸡
Heat shock protein 70Expression in E. coliMonoclonal antibodyWestern-blotImmunohistochemistryBroiler
王志亮,吴晓东,鲍恩东,刘雨田,孙培明,赵永刚.肉鸡HSP70基因克隆、原核表达及单克隆抗体制备[EB/OL].(2006-06-07)[2025-08-11].http://www.paper.edu.cn/releasepaper/content/200606-117.点此复制
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