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目的构建含有乙型肝炎病毒(HBV)S和前S1(preS1,10～50AA)表位的ss1融合基因,并在P.pastoris酵母中表达SS1融合蛋白。方法以HBV全基因组质粒为模板,扩增出目的区段:S(1～222AA)、preS1(10～50AA),以酶切-PCR的方法将其连接为ss1融合基因,然后克隆入表达载体pPIC3.5k;电穿孔转化毕赤酵母菌株GS115,筛选后进行诱导表达并对表达产物进行检测。结果表达产物的相对分子质量为30000左右,与抗HBs抗体和抗preS1抗体均有特异反应。结论ss1融合基因能够在P.pastoris酵母中高效表达,而且表达产物具有HBVS蛋白和preS1蛋白的抗原性,为进一步研究其免疫原性打下基础。
Objective To construct the ss1 fusion gene containing hepatitis B virus (HBV) S and preS1,10 ~ 50AA epitope and express SS1 fusion protein in P. pastoris yeast. Methods The whole genome of HBV was used as a template to amplify the target region: S (1 ~ 222AA) and preS1 (10 ~ 50AA). The recombinant plasmid was ligated into the ss1 fusion gene by restriction enzyme-PCR and cloned into the expression vector pPIC3.5k was transformed into Pichia pastoris strain GS115 by electroporation. After screening, the expression product was induced and the expression product was detected. Results The relative molecular mass of the expressed product was about 30,000, which showed specific reaction with anti-HBs and anti-preS1 antibodies. Conclusion The ss1 fusion gene can be highly expressed in P. pastoris yeast, and the expressed product has the antigenicity of HBVS protein and preS1 protein, which lays the foundation for further study on its immunogenicity.