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Related Experiment Videos

Human papillomavirus type 16 variant assignment by pyrosequencing.

David C Swan1, Josef R Limor, Kara L Duncan

  • 1Division of Viral and Rickettsial Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Public Health Service, U.S. Department of Health and Human Services, Atlanta, GA 30333, USA. dswan@cdc.gov

Journal of Virological Methods
|June 21, 2006
PubMed
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Pyrosequencing offers a faster method for identifying human papillomavirus type 16 (HPV16) variants. This advanced technique accurately detects HPV16 variants, including mixed infections, improving upon traditional sequencing methods.

Area of Science:

  • Virology
  • Molecular Biology
  • Genetics

Background:

  • Human papillomavirus type 16 (HPV16) harbors genetic variations, termed variants, classified geographically (European, African, Asian, Asian-American).
  • HPV16 variants are crucial for distinguishing persistent infections from reinfections and have been linked to varying risks of cervical disease.
  • Traditional variant detection relies on Sanger sequencing, which can be time-consuming.

Purpose of the Study:

  • To evaluate pyrosequencing as a rapid and simultaneous method for detecting HPV16 variants.
  • To compare pyrosequencing results with conventional Sanger sequencing for HPV16 variant determination.

Main Methods:

  • Pyrosequencing was employed to analyze a 314-bp fragment of the HPV16 E6 gene.
  • Biotinylated primers were used for single-stranded template preparation and strand separation.

Related Experiment Videos

  • Seven polymorphic sites (nt 109, 131, 132, 143, 145, 178, 350) were analyzed, including a multiplex pyrosequencing reaction.

    • Main Results:

    • Pyrosequencing successfully identified HPV16 variants in 97 exfoliated cervical samples, confirming Sanger sequencing results.
    • The pyrosequencing method demonstrated high accuracy and efficiency in variant determination.
    • Pyrosequencing detected additional variants in samples with mixed infections, which may have been missed by Sanger sequencing.

    Conclusions:

    • Pyrosequencing is a rapid and effective tool for simultaneous HPV16 variant detection.
    • This method enhances the ability to identify complex HPV16 variant profiles, including mixed infections.
    • Pyrosequencing offers a valuable alternative to conventional sequencing for HPV16 variant analysis in research and clinical settings.