Position: Home > Articles > Cloning and expression of the immunogenic fragment of VP2 protein of canine parvovirus isolated from Nanjing area
Animal Husbandry & Veterinary Medicine
2008,40
(12)
15-18
犬细小病毒南京株VP2蛋白免疫原性片段的克隆表达
作 者：
李希阳;缪勤;张爱华;张洪英;张汇东;张海彬
单 位：
南京农业大学动物医学院;公安部南京警犬研究所;南京市疾病预防控制中心
关键词：
犬细小病毒南京株;VP2基因片段;克隆;表达
摘 要：
以Protean软件对犬细小病毒南京株(CPV-GN)VP2蛋白的分子结构进行分析,从中筛选出具有良好免疫原性潜能的部分片段(VP2′片段),利用PrimerPremier5.0软件设计一对引物,用以该片段的扩增。将PCR扩增产物克隆入pMD18-T载体,构建了pMD-VP2′重组质粒。双酶切回收目的条带,将之亚克隆入表达载体pET32a(+),构建了重组表达质粒pET32-VP2′。转化宿主菌BL21,经IPTG诱导后成功表达了分子量约为34ku的融合蛋白。免疫转印显示,目的蛋白可被CPV-2b抗血清所识别,证明其具有与特异性抗体结合的抗原性,为CPV亚单位疫苗和免疫学诊断方法的研究打下基础。
译 名：
Cloning and expression of the immunogenic fragment of VP2 protein of canine parvovirus isolated from Nanjing area
作 者：
LI Xi-yang1,MIAO Qin2,ZHANG Ai-hua1,ZHANG Hong-ying1,ZHANG Hui-dong2,ZHANG Hai-bin1(1.College of Veterinary Medicine,Nanjing Agricultural University,Nanjing 210095,China;2.Nanjing Policedog Research Institute,Nanjing 210012,China;3.NanJing Center For Disease Control & Prevention,Nanjing 210003,China)
关键词：
CPV-GN;VP2 gene fragment;clone,expression
摘 要：
The molecular structure of VP2 protein of canine parvovirus isolated from Nanjing area(CPV-GN)was predicted by bioinformatic software,from which a highly immunogenic fragment VP2'was screened and amplified by PCR.The PCR product was inserted into pMD 18-T vector and the resulting recombinant plasmid was double digested with EcoR I and Sal I.The expression vector pET32-VP2',designed to propagate and express the fusion protein in Escherichia coli BL21,was constructed by subcloning VP2' into pET32a(+)vector.The desired fusion protein with a molecular weight of 34 ku was detected by SDS-PAGE after induction by IPTG.Subsequent Western bloting assay showed that the recombinant protein could be recognized by the antibody against CPV-2b,and thus has retained the major epitopes of the native protein VP2.
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