水貂阿留申病毒VP2蛋白主要抗原表位基因原核表达及其检测应用

Prokaryotic expression and detective application of the main antigenic regionof VP2 protein of Aleutian Mink Disease Parvovirus

摘要:

将构建好的重组质粒pMD-VP2a、pMD-VP2b分别经双酶切获得基因片段VP2a、VP2b,分别将其定向插入到原核表达载体pMAL-c2中,构建原核表达载体pMAL-VPa和pMAL-VPb。阳性重组质粒转化宿主菌TB1,经IPTG诱导,两段基因均获得表达;Westernblot分析表明表达蛋白具有良好的抗原性。用KCI染色后,切胶回收的方法对表达蛋白进行纯化,以纯化的目的蛋白作为诊断抗原初步建立了水貂阿留申的VP2-CIEP诊断方法。结果表明该方法具有较高的灵敏性和特异性,与传统的CIEP方法的检出符合率达94.3%。

Abstract:

To research safer diagnosis antigen for ADV, the main antigenic region VP2a and VP2b gene of ADV were obtained by restriction digestion of the recombinant plasmids pMD-VP2a and pMD-VP2b. Then the genes were respectively cloned into pMAL-c2 to get two prokaryotic recombinant plasmids pMAL-VPa and pMAL-VPb. The target genes were successfully expressed in the host cell TB1 when induced by IPTG. The Western blot analysis proved the recombinant proteins have good antigenic. The recombinant proteins were purified by KCL dyeing method, and were used as antigen to establish VP2-CIEP for AD diagnoses. The detection result shared 94.3% identity with that of CIEP. The results reported here show that VP2-CIEP is highly sensitive and specific and can benefit the research on the serodiagnosis to AD.