title: A novel accurate amplification created restriction site method for determination of the wild type and the precore mutant hepatitis B virus variants.
abstract: The most commonly occurring hepatitis B virus (HBV) mutation is the G to A mutation at nucleotide 1896 in the precore region. The aim of this study was to develop a novel accurate amplification created restriction site (ACRS) method for determination of the TGG wild type and the TAG precore mutant HBV variants. Two conserved and consensus specific and diagnostic primers introducing BstXI and XagI cleavage sites were designed in order to determine the G1896 wild type and the A1896 precore mutant HBV variants in all HBV genotypes. The results of the ACRS method were compared with sequencing data. With the ACRS method, three different patterns could be distinguished for the wild type, the precore mutant and mixed infection HBV variants. The results of the ACRS method on 30 HBV isolates revealed the TAG precore mutant in 50% (15/30), the TGG wild type variant in 30% (9/30) and the mixed infection in 20% (6/30). The sequencing data of these samples were in agreement with the ACRS results. The ACRS method is a rapid and cost-effective technique for detecting both the TGG wild type and the TAG HBV precore mutant variants. It can be carried out for follow-up of G1896A precore mutant variant in hepatitis B virus infected subjects at routine molecular diagnostic laboratories.
authors.affiliation: Biotechnology Department, Pasteur Institute of Iran, Pasteur Square, Tehran 13164, Iran.
authors.forename: Samad
authors.initials: S
authors.lastname: Amini-Bavil-Olyaee
authors.forename: Ramin
authors.initials: R
authors.lastname: Sarrami-Forooshani
authors.forename: Ahmad
authors.initials: A
authors.lastname: Adeli
authors.forename: Fereidoun
authors.initials: F
authors.lastname: Mahboudi
authors.forename: Farzaneh
authors.initials: F
authors.lastname: Sabahi
authors.forename: Houman
authors.initials: H
authors.lastname: Nafisi
authors.forename: Mohammad Reza
authors.initials: MR
authors.lastname: Zali
authors.forename: Mohammad
authors.initials: M
authors.lastname: Azizi
mesh_terms: D000293:Adolescent; D000328:Adult; D002648:Child; D017931:DNA Primers; D004262:DNA Restriction Enzymes; D005260:Female; D006513:Hepatitis B e Antigens / Q000235:genetics; D006515:Hepatitis B virus / Q000235:genetics / Q000302:isolation & purification*; D019694:Hepatitis B, Chronic / Q000097:blood / Q000453:epidemiology / Q000821:virology*; D006801:Humans; D007492:Iran / Q000453:epidemiology; D008297:Male; D008875:Middle Aged; D009154:Mutation; D009842:Oligopeptides / Q000097:blood / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000191:economics / Q000379:methods*; D012680:Sensitivity and Specificity
chemical_list: D017931:DNA Primers; D006513:Hepatitis B e Antigens; D009842:Oligopeptides; C044757:tyrosyl-glycyl-glycine; C037841:tyrosyl-alanyl-glycine; D004262:DNA Restriction Enzymes