利用穿梭质粒快速构建含乙型肝炎病毒表面抗原(HBsAg)基因的重组杆状病毒  

Rapid construction of recombinant baculovirus carrying hepatitis B virus surface antigen gene by using a novel baculovirus shuttle vector(Bacmid)

作　　者：苗季[1] 丛旭[1] 谭文杰[1] 丛郁[1] 陈刚[1] 詹美云[1] 

机构地区：[1]中国预防医学科学院病毒学研究所肝炎室

出　　处：《中华实验和临床病毒学杂志》1998年第3期245-248,共4页Chinese Journal of Experimental and Clinical Virology

摘　　要：目的应用新型杆状病毒表达系统快速构建含有ＨＢｓＡｇ基因的重组杆状病毒，高效表达ＨＢｓＡｇ，为ＨＢＶ诊断试剂、疫苗及治疗研究提供依据。方法构建含有ＨＢｓＡｇ基因的供体质粒ｐＦＢ－ＢＳ，转化Ｂａｃ－ｔｏ－Ｂａｃ杆状病毒表达试剂盒中的ＤＨ１０Ｂａｃ致敏菌，利用其含有的细菌Ｔｎ７转座系统将ＨＢｓＡｇ基因重组至穿梭质粒Ｂａｃｍｉｄ上，快速筛选出含有ＨＢｓＡｇ基因的重组杆状病毒。结果此重组病毒能在昆虫细胞中表达出有生物学活性的ＨＢｓＡｇ，表达产物主要集中在细胞内，ＥＬＩＳＡ滴度为１：２×１０４，产量可以达到２μｇ／１０６细胞。In order to express the HBV surface antigen (HBV) efficiently by using the Bac-to-Bac system and to produce HBsAg for diagnosis and vaccine development of HBV, a recombinant donor plasmid pFB-BS carrying HBsAg gene was constructed. The gene was inserted into shuttle plasmid (Bacmid) in E.coli DH10Bac with the help of Tn7 transposition system. The recombinant Bacmid was examined by PCR and transfected to Sf9 cells. HBsAg was expressed under the control of polyhedrin promoter of baculovirus in insect cells and was identified by ELISA. The quantity of the expression reached to 2μg/10 6 cells and most production was within cells. Our data suggested that the Bac-to-Bac system can be used successfully and rapidly in the expression of viral gene.

关 键 词：杆状病毒 蛋白表达 乙型肝炎病毒 表面抗原 

分 类 号：R373.21[医药卫生—病原生物学]