| 诊断用丁肝抗原的基因优化及其蛋白表达、纯化和鉴定 |
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| 引用本文： | 丁军颖,郭敏卓,伊瑶,苏秋东,卢学新,邱丰,毕胜利.诊断用丁肝抗原的基因优化及其蛋白表达、纯化和鉴定[J].中华实验和临床病毒学杂志,2012,26(2):87-89. |
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| 作者姓名： | 丁军颖 郭敏卓 伊瑶 苏秋东 卢学新 邱丰 毕胜利 |
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| 作者单位： | 1. 中国疾病预防控制中心病毒病预防控制所,北京,102206
2. 北京出入境检验检疫局保健中心 |
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| 基金项目： | 国家科技重大专项（2008ZX10201） |
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| 摘 要： | 目的 获取有生物活性的丁肝抗原蛋白,探讨其作为诊断试剂的应用价值.方法 经密码子优化,合成人工编码的丁肝抗原基因序列；以M48作为表达载体,构建带有硫氧还蛋白的重组表达质粒；在大肠埃希氏菌中经IPTG诱导表达；以亲和层析纯化并以ELISA鉴定该蛋白.结果 酶切鉴定构建的质粒正确；SDS-PAGE结果显示表达和纯化的外源蛋白与预期相对分子质量大小一致；ELISA鉴定该蛋白有与丁肝抗体特异性结合的活性.结论 成功获得有生物活性的可供诊断用的丁肝抗原蛋白,为丁肝诊断试剂的研发奠定了基础.
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| 关 键 词： | 基因 硫氧还蛋白 色普法 肝炎 丁型 |
Gene optimization, protein expression, purification and characterization of the HDV antigen for diagnosis |
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| DING Jun-ying
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GUO Min-zhuo
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YI Yao
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SU Qiu-dong
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LU Xue-xin
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QIU Feng
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BI Sheng-li.Gene optimization, protein expression, purification and characterization of the HDV antigen for diagnosis[J].Chinese Journal of Experimental and Clinical Virology,2012,26(2):87-89. |
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| Authors: | DING Jun-ying GUO Min-zhuo YI Yao SU Qiu-dong LU Xue-xin QIU Feng BI Sheng-li |
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| Institution: | . National Institute For Viral Disease Control and Prevention ,Beijng 102206, China |
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| Abstract: | Objective To prepare HDAg with biological activities as a candidate of diagnostic reagent. Methods To synthesize HDV gene fragment after codon optimization. To construct a thio-fused recombinant plasmid based on M48 expression vector. To express in E. coli induced by IPTG. To purify the protein by affinity chromatography followed by characterization in ELISA. Results Plasmid construction was verified by enzyme digestion. SDS-PAGE indicated the molecular weight of the protein was the same as we expectation. ELISA proved its affinity with HDV antibodies. Conclusion HDAg was obtained successfully and it will pave the road to the research of HDV diagnostic reagent. |
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| Keywords: | Genes Thioredoxim Chromatography Hepatitis D |
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