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当前位置: 首页 > 文章 > 猪戊型肝炎病毒ORF2蛋白N端主要抗原表位区的原核表达 中国畜牧兽医 2010,31 (1) 54-57
Position: Home > Articles > Expression of N-terminal Major Epitope Domain of ORF2 of SHEV in E. coli China Animal Husbandry & Veterinary Medicine 2010,31 (1) 54-57

猪戊型肝炎病毒ORF2蛋白N端主要抗原表位区的原核表达

作  者：
冉旭华;闻晓波;王密;李冬野;侯喜林
单  位：
黑龙江八一农垦大学动物科技学院预防兽医学省重点实验室
关键词：
猪戊型肝炎病毒;ORF2;主要抗原表位区;表达
摘  要：
用原核表达系统表达猪戊型肝炎病毒ORF2蛋白N端的主要抗原表位区。通过对GenBank公布的猪戊型肝炎病毒DQ1株ORF2的氨基酸序列进行分析,确定了其N端抗原性较高的区域,针对此区域设计并合成了1对特异性引物,RT-PCR扩增该区段,并将此片段克隆到原核表达载体pET30a(+)上,命名为pET30a-ORF2N,转化大肠杆菌BL21感受态细胞,1.0 mmol/LIPTG37℃诱导表达。RT-PCR扩增得到424 bp的片段,重组蛋白大小约为21.8 ku,与预期大小相符。West-ern blotting分析结果表明,该蛋白能与阳性血清发生特异性反应,证明该蛋白具有生物学活性。猪戊型肝炎病毒ORF2蛋白N端主要抗原表位区在大肠杆菌中成功表达,具有一定的反应原性,为进一步用其建立诊断方法奠定基础。
译  名：
Expression of N-terminal Major Epitope Domain of ORF2 of SHEV in E. coli
作  者：
RAN Xu-hua,WEN Xiao-bo,WANG Mi,LI Dong-ye,HOU Xi-lin(Provincial Key Laboratory of Preventive Veterinary Medicine,College of Animal Science and Technology,Heilongjiang Bayi Agricultural University,Daqing 163319,China)
关键词：
swine hepatitis E;ORF2;major epitope domain;expression
摘  要：
To express N-terminal major epitope domain of ORF2 of swine hepatitis E(SHEV) in E.coli.According to the amino acid sequence of SHEV DQ1 strain in GenBank,N-terminal major epitope domain in ORF2 was primarily mapped upon analysis of bioinformatic software.A pair of specific primer was designed to amplify the fragment encoding the major epitope domain.This fragment was cloned into the multiple cloning sites of pET30a(+) vector after subjecting to restriction endonuclease digestion.The recombinant plasmid was designated pET30a-ORF2N and was transformed into E.coli BL21 competent cells.Expression of recombinant protein was induced by addition of final concentration of 1.0 mmol/L IPTG under 37 ℃ condition.The length of RT-PCR product of interest is 424 base pairs.The size of recombinant amino acid deduced is 21.8 ku in theory.The result of Western blotting showed that the recombinant protein specifically reacted with serum against SHEV,suggesting the recombinant protein had wonderful specificity.N-terminal major epitope domain of ORF2 of SHEV was expressed successfully in E.coli and showed specificity.It can be used as antigen in diagnostic assay for measuring antibodies against SHEV in clinical practice.

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