文章摘要
尹全章,姚 堃,王 芳,许文嵘,徐 建,陈 云,周 锋,冯东举.人类疱疹病毒6型U94基因克隆?表达?纯化及抗体制备[J].南京医科大学学报,2007,(8):847~851
人类疱疹病毒6型U94基因克隆?表达?纯化及抗体制备
Cloning,expression and purification of human herpesvirus 6 U94 and its antibodies production
投稿时间:2007-04-20  
DOI:10.7655
中文关键词: 人类疱疹病毒6型  U94  基因表达  抗体,病毒
英文关键词: HHV6  U94  gene expression  antibodies,viral
基金项目:江苏省自然科学基金资助项目(BK97051)
作者单位
尹全章 南京医科大学微生物与免疫学系,江苏 南京 210029 
姚 堃  
王 芳  
许文嵘  
徐 建  
陈 云  
周 锋  
冯东举  
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中文摘要:
      目的:制备人类疱疹病毒6型U94蛋白及其抗体?方法:PCR扩增U94基因,经测序后克隆到原核表达载体PGEX-6p-1中?重组质粒转化大肠杆菌Rosetta,诱导蛋白表达?应用酶切鉴定?SDS-PAGE及Western blot等方法确保基因片段的正确性及表达蛋白的特异性?表达的蛋白经亲和层析纯化,纯化的蛋白免疫动物制备抗血清?结果:成功地获得了高纯度的U94融合蛋白,纯度可达90%以上?用其制备多克隆抗体滴度可达1:100 000?结论:获得高纯度的U94融合蛋白及其高效价的抗体?
英文摘要:
       Objective:To prepare human herpesvirus 6 U94 protein and produce its antibodies. Methods:U94 gene of HHV6 was amplified by PCR and sequenced. The resulting DNA construct was cloned into a prokaryotic expression vector(PGEX-6P-1). The recombinant plasmid was transformed into Eserichia coli Rosetta. The accuracy of inserted gene and specificity of proteins were detected by two enzymes digestion, SDS-PAGE, and Western Blot. The fusion U94 protein purified by affinity chromatograph was used to vaccinate rabbit to produce antibodies. Results:The purity of U94 protein was above 90%. The titer of the antibodies was 1 ∶ 100 000. Conclusion:HHV6 U94 fusion protein with high purity was obtained and its corresponding polyclonal antibodies with high titer were produced.
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