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Molecular Diagnosis & Therapy

, Volume 10, Issue 2, pp 125–134 | Cite as

Simultaneous Detection of Precore/Basal Core Promoter Mutations in Hepatitis B Virus Using Arrayed Primer Extension

  • Wai-Yan Ha
  • Chi-Chiu Lau
  • Patrick Y. K. Yue
  • Kaman K. M. Hung
  • Kelvin Chan
  • Siu-Hon Chui
  • Albert K. K. Chui
  • Wing-Cheong Yam
  • Ricky N. S. Wong
Original Research Article

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.

Keywords

Core Promoter Basal Core Promoter Core Promoter Region Core Promoter Mutation HBeAg Negativity 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Acknowledgments

The first two authors contributed equally to this work.

This work was supported by a grant from Vigconic (International) Limited.

The academic institutions or employers involved in this manuscript do not have any financial interest in or financial conflict with the subject matter or materials discussed in this manuscript.

The authors have no conflicts of interest that are directly relevant to the content of this article.

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Copyright information

© Adis Data Information BV 2006

Authors and Affiliations

  • Wai-Yan Ha
    • 1
  • Chi-Chiu Lau
    • 1
  • Patrick Y. K. Yue
    • 1
  • Kaman K. M. Hung
    • 1
  • Kelvin Chan
    • 2
  • Siu-Hon Chui
    • 3
  • Albert K. K. Chui
    • 4
  • Wing-Cheong Yam
    • 5
  • Ricky N. S. Wong
    • 1
    • 6
  1. 1.Research and Development Division, School of Chinese MedicineHong Kong Baptist UniversityHong Kong
  2. 2.Department of Pharmacy, School of Applied SciencesUniversity of WolverhamptonWolverhamptonUK
  3. 3.Diagnostix Medical Centre LtdHong Kong
  4. 4.Department of SurgeryThe Chinese University of Hong Kong, Prince of Wales HospitalHong Kong
  5. 5.Department of Microbiology, Faculty of MedicineThe University of Hong KongHong Kong
  6. 6.Department of Biology, Science FacultyHong Kong Baptist UniversityKowloon TongHong Kong

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