title: Generation of Replication-Competent Hepatitis B Virus Genome from Blood Samples for Functional Characterization.
abstract: Hepatitis B virus (HBV) infection can be associated with a spectrum of clinical outcomes. Transient transfection of the clinical HBV isolates in human hepatoma cell lines can establish their biological properties to shed light on their different pathogenic potentials, yet very few clinical HBV isolates have been functionally characterized so far. The technical challenges include faithful amplification of the full-length HBV genome from clinical samples and conversion into a replication-competent form. We have improved a published method to amplify the full-length HBV genome from blood samples. Two alternative approaches are used to render the cloned HBV genome replication competent: release and circularization of the 3.2-kb HBV genome prior to each transfection experiment or conversion of the monomeric clone into a tandem dimer version.
authors.affiliation: Liver Research Center, Rhode Island Hospital, Brown University, Providence, RI, USA.|Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai, China.
authors.forename: Yanli
authors.initials: Y
authors.lastname: Qin
authors.affiliation: Department of Pathogen Biology, School of Basic Medical Sciences, Fudan University, Shanghai, China.
authors.forename: Yong-Xiang
authors.initials: YX
authors.lastname: Wang
authors.affiliation: Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai, China.
authors.forename: Jiming
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Liver Research Center, Rhode Island Hospital, Brown University, Providence, RI, USA.
authors.forename: Jisu
authors.initials: J
authors.lastname: Li
authors.affiliation: Liver Research Center, Rhode Island Hospital, Brown University, Providence, RI, USA. shuping_tong_md@brown.edu.|Department of Pathogen Biology, School of Basic Medical Sciences, Fudan University, Shanghai, China. shuping_tong_md@brown.edu.
authors.forename: Shuping
authors.initials: S
authors.lastname: Tong
mesh_terms: D002460:Cell Line; D003001:Cloning, Molecular; D004270:DNA, Circular; D004279:DNA, Viral; D016679:Genome, Viral; D006509:Hepatitis B / Q000821:virology*; D006515:Hepatitis B virus / Q000302:isolation & purification / Q000502:physiology*; D006801:Humans; D016133:Polymerase Chain Reaction; D012367:RNA, Viral; D014158:Transcription, Genetic; D014162:Transfection; D014771:Virion; D014779:Virus Replication / Q000235:genetics*
chemical_list: D004270:DNA, Circular; D004279:DNA, Viral; D012367:RNA, Viral
keywords: HBV clinical isolates; Tandem dimer; Transfection