【摘要】 目的构建艰难梭菌(CD)谷氨酸脱氢酶(GDH)的原核表达载体,表达重组蛋白并鉴定其抗原活性。方法以CD的ATCC43255菌株基因组DNA为模板,通过PCR法扩增GDH全长基因片段,构建原核表达载体并转化到大肠杆菌中,IPTG诱导表达GDH蛋白,并用Ni柱进行纯化。利用CD抗原检测试剂盒对GDH抗原区段的抗原性进行鉴定。结果构建的原核表达载体p ET-28a/GDH可在大肠杆菌中成功表达,获得的GDH抗原能与相应抗体产生特异性反应。结论原核表达的CD的GDH抗原具有很好的抗原活性,为进一步制备相应抗体并建立CD的GDH抗原的免疫检测方法奠定了基础。更多还原

【Abstract】 Objective To construct prokaryotic expression vectors for glutamate dehydrogenase( GDH) of Clostridium difficile( CD),and express recombinant GDH in Escherichia coli,and identify its antigenicityed. Methods The entire gene of GDH was cloned from ATCC43255 genome DNA. The recombinant antigens were expressed in E. coli with IPTG induction and purified by Ni-NATBeads. The antigenicity was detected using CD Qick Chek Complete dual-antigen EIA.Results Prokaryotic expression vectors of CD GDH were constructed successfully. The antigen could be identified by specific anti-GDH antibodies. Conclusion The GDH antigen can be used to prepare corresponding antibodies,which facilitate the development of immunoassay for CD GDH.更多还原