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    Simultaneous detection of precore/basal core promoter mutations in hepatitis B virus using arrayed primer extension.

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    Authors
    Ha, Wai-Yan
    Lau, Chi-Chiu
    Yue, Patrick Y. K.
    Hung, Kaman K. M.
    Chan, Kelvin C.
    Chui, Siu-Hon
    Chui, Albert K. K.
    Yam, Wing-Cheong
    Wong, Ricky N. S.
    Issue Date
    2006
    Submitted date
    2007-02-08
    
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    Abstract
    BACKGROUND: Hepatitis B is a major disease that causes serious public health problems worldwide. The loss of HBeAg expression due to point mutations or single nucleotide polymorphisms (SNPs) in the precore/basal core promoter region of the hepatitis B virus (HBV) is associated with hepatocellular cirrhosis and carcinoma. Simultaneous screening for these mutations is strongly advocated for monitoring disease development in HBV-infected patients. The aim of this study is to apply arrayed primer extension (APEX) for the detection of HBV SNPs at the precore/basal core promoter. METHODS AND RESULTS: We optimized APEX for simultaneous detection of eight potential sites of SNPs in the precore/basal core promoter region of HBV. The precore/basal core promoter regions of HBV from 36 HBV-infected patients were amplified by PCR. After purification and DNA fragmentation, the short, single-stranded HBV DNA fragments were allowed to hybridize with the oligonucleotides corresponding to the sites of SNPs immobilized on glass slides, followed by incorporation of different fluorescently labeled dideoxynucleotides. This allows fast and unequivocal discrimination between wild-type and mutant genotypes with high dideoxy-nucleotide incorporation efficiency, sensitivity, and specificity. The coexistence of both genotypes was also detected; this was undetected by DNA sequencing. CONCLUSION: The simultaneous detection of SNPs in HBV precore/basal core promoter by APEX enables large-scale diagnostic analysis, which can be extended to the whole HBV genome.
    Citation
    Molecular Diagnosis & Therapy, 10(2): 125-134
    Publisher
    Adis
    URI
    http://hdl.handle.net/2436/15153
    PubMed ID
    16669611
    Additional Links
    http://direct.bl.uk/bld/PlaceOrder.do?UIN=188017433&ETOC=RN&from=searchenginehttp://www.ingentaconnect.com/content/adis/mdt/2006/00000010/00000002/art00007
    Type
    Journal article
    Language
    en
    Description
    Metadata only
    ISSN
    1177-1062
    Collections
    Research Institute in Healthcare Science

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