人IL-2与HBsAg融合基因重组卡介苗的构建与鉴定  

Construction and identification of recombinant BCG with human IL-2 and HBsAg fusion gene

作　　者：王子豪 于沛伶 潘玉成 李运清 刘昂 薛庆节 Wang Zihao;Yu Peiling;Pan Yucheng;Li Yunqing;Liu Ang;Xue Qingjie(Basic Medical College,Jining Medical University,Jining 272067,China)

机构地区：[1]济宁医学院基础医学院,272067

出　　处：《国际生物制品学杂志》2021年第1期28-32,共5页International Journal of Biologicals

基　　金：山东省重点研发计划项目(2018GSF118137);山东省医药卫生科技发展计划项目(2017WS339);山东省高等学校科技计划项目(J17KB085);济宁市重点研发计划项目(2019SMNS020);济宁医学院重点项目(16008);贺林院士工作站重点项目(JYHL2019ZD03);济宁医学院大创项目(CX2018007,CX2019102)。

摘　　要：目的构建人IL-2与HBsAg融合基因并转化到BCG中构建重组BCG(recombinant BCG,rBCG),为制备治疗和预防乙型肝炎和结核病的二联疫苗奠定基础。方法以IL-2 pMalLP质粒和HBsAg pMalLP质粒为模板,设计引物,利用PCR技术扩增HBsAg和IL-2基因,进行重叠PCR获得融合基因IL-2-HBsAg并纯化,将融合基因克隆至穿梭表达载体pMV261得到重组质粒,电转化导入BCG感受态细胞,以蛋白质印迹法检测融合基因在rBCG中的表达。结果 PCR扩增得到IL-2和HBsAg,通过重叠PCR获得IL-2-HBsAg融合基因后成功插入穿梭表达载体,rBCG能表达融合蛋白。结论构建人IL-2与HBsAg融合基因重组质粒并导入BCG,获得了稳定表达融合蛋白的rBCG。Objective To construct the fusion gene of human IL-2 and HBsAg and transform into BCG for expression,laying foundation for preparation of double vaccine for prevention and treatment of hepatitis B and tuberculosis.Methods Using IL-2 pMalLP and HBsAg pMalLP as templates,primers were designed.HBsAg and IL-2 genes were amplified by PCR,and then the fusion gene IL-2-HBsAg was obtained by overlapping PCR and purified.The fusion gene was cloned on the shuttle expression vector pMV261 to obtain the recombinant plasmid,and then the fusion gene was transformed into BCG sensitive cells by electricity,and the expression of the fusion gene in recombinant BCG(rBCG)was detected by Western blot.Results IL-2(435 bp)gene and HBsAg(681 bp)gene were amplified by PCR,and IL-2-HBsAg(1179 bp)fusion gene was amplified by overlapping PCR.The fusion gene was successfully inserted into the shuttle expression vector.The fusion protein was detected in rBCG.Conclusion The recombinant plasmid of human IL-2 and HBsAg fusion gene is successfully constructed and introduced into BCG to obtain rBCG which can stably express the fusion protein.

关 键 词：白细胞介素2 肝炎病毒 乙型 重组融合蛋白质类卡介苗 肝炎表面抗原 乙型 

分 类 号：R373.1[医药卫生—病原生物学]