Citation: W ANG Han—zhong, HUANG Yi, SI Yan—hong, FANG Ming.gang . CHEN Xinwen, Just M .Vlak .HU Zhi.hong. A Novel Expression System of Helicoverpa armigera single-nucleocapsid nucleopolydrovirus .VIROLOGICA SINICA, 2002, 17(4) : 319-325.

A Novel Expression System of Helicoverpa armigera single-nucleocapsid nucleopolydrovirus

  • Available online: 05 November 2002
  • A 8.5kb fragment containing an E .coli low—copy number mini F replicon,a selectable kanamycin resistance marker and lacZ gene with attTn7(the target site for bacterial transposon Tn7) replaced polyhedrin gene in HaSNPV genome using homologous recombination.HaSNPV genome was cloned and maintained as 1 32 kb bacterial artificial chromosome(Bacmid)in E.coli,transfection of the Bacmid(HaBacmid.HZ8)into HzAml cells led to a productive virus infection.In this paper.the donor plasmid HapFastPhP10 was constructed using the Ha—polh gene and the Ha.P10 promoter tO re. place the original Ac P1 0 promoter and Ac—Polh promoter of the pFastBacDual donor plasmid respec. tively.The eGFP gene was inserted into multiple cloning site(MCS)downsream of the P10 promoter in the pFastBacHaPhpP10 donor plasmid.Recombinant HaBacmid HZ8 was constructed by transpo s. ing a mini—Tn7 element from a HaDFastPhP1 0 donor plasmid tO the mini.attTn7 attachment site on the HaBacmid—HZ8 when the Tn7 transposition functions are provided in trans by a helper plasmid.Re. combinant HaBacmid.HZ8 DNA was transfected HzAml cells.occlusion bodies and green flusecent were found within HzAml cells 5 days after transfection.The results indicated the Habac to Bac ex. pression system based on HaSNPV can effectively express foreign gene.

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    A Novel Expression System of Helicoverpa armigera single-nucleocapsid nucleopolydrovirus

    • 1. 1.Joint—lab ofInvertebrate I/Trology,Wuhan Instituteof Virology,ChineseAcademyof Science,Wuhan 430071,China: 2.Labboratoryof Virology,Wageningen University,6709 PD 讹geningen,TheNetherlands

    Abstract: A 8.5kb fragment containing an E .coli low—copy number mini F replicon,a selectable kanamycin resistance marker and lacZ gene with attTn7(the target site for bacterial transposon Tn7) replaced polyhedrin gene in HaSNPV genome using homologous recombination.HaSNPV genome was cloned and maintained as 1 32 kb bacterial artificial chromosome(Bacmid)in E.coli,transfection of the Bacmid(HaBacmid.HZ8)into HzAml cells led to a productive virus infection.In this paper.the donor plasmid HapFastPhP10 was constructed using the Ha—polh gene and the Ha.P10 promoter tO re. place the original Ac P1 0 promoter and Ac—Polh promoter of the pFastBacDual donor plasmid respec. tively.The eGFP gene was inserted into multiple cloning site(MCS)downsream of the P10 promoter in the pFastBacHaPhpP10 donor plasmid.Recombinant HaBacmid HZ8 was constructed by transpo s. ing a mini—Tn7 element from a HaDFastPhP1 0 donor plasmid tO the mini.attTn7 attachment site on the HaBacmid—HZ8 when the Tn7 transposition functions are provided in trans by a helper plasmid.Re. combinant HaBacmid.HZ8 DNA was transfected HzAml cells.occlusion bodies and green flusecent were found within HzAml cells 5 days after transfection.The results indicated the Habac to Bac ex. pression system based on HaSNPV can effectively express foreign gene.

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