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Virus Genome Detection by the Polymerase Chain Reaction (PCR)

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Practical Molecular Virology

Part of the book series: Methods in Molecular Biology ((MIMB,volume 8))

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Abstract

Since the polymerase chain reaction (PCR) technique was developed in 1986 (1), it has found wide application in molecular biology and is now regarded as an irreplaceable tool in many laboratories. This chapter will deal with its use for the detection of virus genomes for diagnostic and experimental purposes.

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References

  1. Saiki, R. K., Gelfand, D. H., Stoffel, S., Scharf, S. J., Higuchi, R, Horn, G. T., Mullis, K. B., and Erlich, H. A. (1988) Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science 239, 487–491.

    Article  PubMed  CAS  Google Scholar 

  2. Goodenow, M., Huet, T., Saurin, W., Kwok, S., Sninsky, J., and Wain-Hobson, S. (1989) HIV-1 isolates are rapidly evolving quasispecies: Evidence for viral mixtures and preferred nucleotide substitutions. J. Acqu. Immunodefic. Syndr. 2, 344–352.

    CAS  Google Scholar 

  3. Sambrook, J., Fritsch, E. F., and Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual, 2nd Ed. (Cold Spring Harbor Laboratory, Cold Spring Harbor, NY).

    Google Scholar 

  4. Rodgers, M. F., Ou, C.-Y, Rayfield, M., Thomas, P. A., Schoenbaum, E. E., Abrams, E., Krasinski, K., Selwyn, P. A., Moore, J., Kaul, A., Grimm, K. T., Bamji, M, and Schochetman, G. (1989) Use of the polymerase chain reaction for early detection of the proviral sequences of human immunodeficiency virus in infants born to seropositive mothers. N. Engl. J. Med. 320, 1649–1654.

    Article  Google Scholar 

  5. Simmonds, P., Balfe, P., Peutherer, J. F., Ludlam, C. A., Bishop, J. O., and Brown, A. J. L. (1990) Human immunodeficiency virus-infected individuals contain provirus in small numbers of peripheral mononuclear cells and at low copy numbers. J. Virol. 64, 864–872.

    PubMed  CAS  Google Scholar 

  6. Abbott, M. A., Poiesz, B. J., Byrne, B. C., Kwok, S., Sninsky, J. J., and Ehrlich, G. D. (1988) Enzymatic gene amplification: Qualitative and quantitative methods for detecting proviral DNA amplified in vitro. J. Infect. Dis 158, 1158–1169.

    PubMed  CAS  Google Scholar 

  7. Ou, C.-Y., Kwok, S., Mitchell, S. W., Mack, D., Sninsky J. J., Krebs, J. W, Feorino, P., Warfield, D., and Schochetman, G. (1988) DNA amplification for direct detection of HIV-1 in DNA of peripheral blood mononuclear cells. Science 239, 295–297

    Article  PubMed  CAS  Google Scholar 

  8. Chen, E. Y. and Seeburg, P. H. (1985) Supercoil sequencing: A fast and simple method for sequencing plasmid DNA. DNA 4(2), 165–170.

    Article  PubMed  CAS  Google Scholar 

  9. Winship, P. R. (1989) An improved method for directly sequencing PCR amplified material using dimethyl sulphoxide. Nucleic Acids Res 17, 1266.

    Article  PubMed  CAS  Google Scholar 

  10. Gyllensten, U. B. and Erlich, H. A. (1988) Generation of single stranded DNA by the polymerase chain reaction and its application to direct sequencing of the HLA-DQA locus. Proc. Nat. Acad. Sa. USA 85, 7652–7656.

    Article  CAS  Google Scholar 

  11. Reddy, P., Sandberg-Wollheim, M., and Mettus, R. (1989) Amplification and molecular cloning of HTLV-1 sequences from DNA of MS patients. Science 243, 529–533

    Article  PubMed  CAS  Google Scholar 

  12. Richardson, J., Wucherpfennig, K, Endo, N., Rudge, P., Dalgleish, A. G., and Hafler, D. A. (1989) PCR analysis of DNA from MS patients for the presence of HTLV-1. Science 246, 821–823.

    Article  CAS  Google Scholar 

  13. Sarkar, G. and Sommer, S S (1990) Shedding light on PCR contamination Nature 343, 27.

    Article  PubMed  CAS  Google Scholar 

  14. Bell, J. and Ratner, L. (1989) Specificity of polymerase chain amplification reactions for human immunodeficiency virus type 1 DNA sequences. AIDS Res Hum. Retroviruses 5, 87–95.

    Article  PubMed  CAS  Google Scholar 

  15. Laure, F., Courgnaud, V., Rouzioux, C., Blanche, S., Veber, F., Burgard, M., Jacomet, C., Griscelli, C., and Brechot, C. (1988) Detection of HTV-1 DNA in infants and children by means of the polymerase chain reaction. Lancet 2, 538–541.

    Article  PubMed  CAS  Google Scholar 

  16. Loche, M. and Mach, B. (1988) Identification of HFV-infected seronegative individuals by a direct diagnostic test based on hybridisation to amplified viral DNA Lancet 2, 418–421.

    Article  PubMed  CAS  Google Scholar 

  17. Ou, C.-Y., Kwok, S., Mitchell, S. W., Mack, D. H., Sninsky, J. J., Krebs, J. W., Feorino, P., Warfield, D., and Schochetman, G. (1988) DNA amplification for direct detection of HIV-1 in DNA of peripheral blood mononuclear cells. Science 239, 295–297.

    Article  PubMed  CAS  Google Scholar 

  18. Arrigo, S. J., Weitsman, S., Rosenblatt, J D, and Chen, I S. (1989) Analysis of rev gene function on human immunodeficiency virus type 1 replication in lymphoid cells by using a quantitative polymerase chain reaction method. J. Virol. 63, 4875–4881

    PubMed  CAS  Google Scholar 

  19. Weber, J., Clapham, P., McKeating, J., Stratton, M., Robey, E., and Weiss, R. (1989) Infection of brain cells by diverse human immunodeficiency virus isolates: Role of CD4 as receptor. J. Gen. Virol 70, 2653–2660.

    Article  PubMed  CAS  Google Scholar 

  20. Hart, C., Schochetman, G., Spira, T., Iifson, A., Moore, J., Galphin, J., Sninsky, J., and OU, C.-Y. (1988) Direct detection of HTV RNA expression in seropositive subjects Lancet 2, 596–599.

    Article  PubMed  CAS  Google Scholar 

  21. Rayfield, M., De-Cock, K., Heyward, W., Goldstein, L., Krebs, J., Kwok, S., Lee, S, McCormick, J., Moreau, J. M., Odehouri, K., Schochetman, G., Sninski, J, and Ou, C-Y. (1988) Mixed human immunodeficiency virus (HIV) infection in an individual. Demonstration of both HIV type 1 and type 2 proviral sequences by using polymerase chain reaction. J. Infect. Dis. 158, 1170–1176.

    PubMed  CAS  Google Scholar 

  22. Lynas, C., Cook, S. D., Laycock, K. A., Bradfield, J. W, and Maitland, N. J. (1989) Detection of latent virus mRNA in tissues using the polymerase chain reaction. J. Pathol. 157, 285–289.

    Article  PubMed  CAS  Google Scholar 

  23. Cao, M., Xiao, X, Egbert, B., Darragh, T M, and Yen, T S (1989) Rapid detection of cutaneous herpes simplex virus infection with the polymerase chain reaction. J. Invest. Dermatol. 92, 391–392.

    Article  PubMed  CAS  Google Scholar 

  24. Gama, R. E., Horsnell, P. R, Hughes, P. J, North, C., Bruce, C. B., al-Nakib, W., and Stanway, G. (1989) Amplification of rhinovirus specific nucleic acids from clinical samples using the polymerase chain reaction. J Med. Virol. 28, 73–77.

    Article  PubMed  CAS  Google Scholar 

  25. Saito, I., Servenius, B., Compton, T., and Fox, R. I. (1989) Detection of Epstein-Barr virus DNA by polymerase chain reaction in blood and tissue biopsies from patients with Sjogren’s syndrome. J. Exp. Med. 169, 2191–2198.

    Article  PubMed  CAS  Google Scholar 

  26. Thiers, V., Nakajima, E., Kremsdorf, D., Mack, D., Schellekens, H., Driss, F., Goudeau, A., Wands, J., Sninsky, J., Tiollais, P., and Brechot, C. (1988) Transmission of hepatitis B from hepatitis-B seronegative subjects. Lancet 2, 1273–1276.

    Article  PubMed  CAS  Google Scholar 

  27. Kaneko, S., Miller, R. H., Feinstone, S. M., Unoura, M., Kobayashi, K., Hattori, N., and Purcell, R. H. (1989) Detection of serum hepatitis B virus DNA in patients with chronic hepatitis using the polymerase chain reaction assay. Proc. Natl Acad. Sci USA 86, 312–316.

    Article  PubMed  CAS  Google Scholar 

  28. Sumazaki, R, Motz, M, Wolf, H., Heinig, J., Jilg, W, and Deinhardt, F. (1989) Detection of hepatitis B virus in serum using amplification of viral DNA by means of the polymerase chain reaction J. Med. Virol 27, 304–308.

    Article  PubMed  CAS  Google Scholar 

  29. Shibata, D., Martin, W. J., Appleman, M. D, Causey, D. M., Leedom, J M., and Amheim, N. (1988) Detection of cytomegalovirus DNA in peripheral blood of patients infected with human immunodeficiency virus. J Infect. Dis. 158, 1185–1192.

    PubMed  CAS  Google Scholar 

  30. Demmler, G. J., Buffone, G. J, Schimbor, C. M, and May, R A (1988) Detection of cytomegalovirus in urine from newborns by using polymerase chain reaction DNA amplification. J. Infect. Dis. 158, 1177–1184.

    PubMed  CAS  Google Scholar 

  31. Young, L. S., Bevan, I. S., Johnson, M. A., Blomfield, P. I, Bromidge, T., Maitland, N. J., and Woodman, C. B. (1989) The polymerase chain reaction: A new epidemiological tool for investigating cervical human papillomavirus infection. Br. Med. J. 298, 14–18.

    Article  CAS  Google Scholar 

  32. Maitland, N. J., Bromidge, T., Cox, M. F., Crane, I. J., Prime, S. S., and Scully, C. (1989) Detection of human papillomavirus genes in human oral tissue biopsies and cultures by polymerase chain reaction. Br.J. Cancer 59, 698–703.

    Article  PubMed  CAS  Google Scholar 

  33. Shibata, D. K., Arnheim, N., and Martin, W.J. (1988) Detection of human papilloma virus in paraffin-embedded tissue using the polymerase chain reaction. J. Exp. Med. 167, 225–230.

    Article  PubMed  CAS  Google Scholar 

  34. Kinoshita, T., Shimoyama, M., Tobinai, K., Ito, M., Ito, S., Ikeda, S., Tajima, K., Shimotohno, K., and Sugimura, T. (1989) Detection of mRNA for the tax1/rex1 gene of human T-cell leukemia virus type I in fresh peripheral blood mononuclear cells of adult T-cell leukemia patients and viral carriers by using the polymerase chain reaction. Proc. Natl Acad. Sci. USA 86, 5620–5624.

    Article  PubMed  CAS  Google Scholar 

  35. Greenberg, S. J., Ehrlich, G. D., Abbott, M. A., Hurwitz, B. J., Waldman, T. A., and Poiesz, B. J. (1989) Detection of sequences homologous to human retroviral DNA in multiple sclerosis by gene amplification. Proc. Natl. Acad. Sci. USA 86, 2878–2882.

    Article  PubMed  CAS  Google Scholar 

  36. Arthur, R. R., Dagostin, S., and Shah, K. V. (1989) Detection of BK virus and JC virus in urine and brain tissue by the polymerase chain reaction. J. Clin. Microbiol. 27, 1174–1179.

    PubMed  CAS  Google Scholar 

  37. Schulz, T. F, Whitby, D., Hoad, J. G., Corrah, T., Whittle, H., and Weiss, R. A Biological and molecular variability of HIV-2 isolates from the Gambia. J. Virol. 64(10), 5177–5182.

    Google Scholar 

  38. Lee, H., Swanson, P., Shorty, V. S., Zack, J. A., Rosenblatt, J. D., and Chen, I S. Y. (1989) High rate of HTLV-II infection in seropositive IV drug abusers in New Orleans. Science 244, 471–475.

    Article  PubMed  CAS  Google Scholar 

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© 1991 The Humana Press Inc., Clifton, NJ

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Hoad, J.G., Schulz, T.F. (1991). Virus Genome Detection by the Polymerase Chain Reaction (PCR). In: Collins, M.K.L. (eds) Practical Molecular Virology. Methods in Molecular Biology, vol 8. Humana Press. https://doi.org/10.1385/0-89603-191-8:61

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  • DOI: https://doi.org/10.1385/0-89603-191-8:61

  • Publisher Name: Humana Press

  • Print ISBN: 978-0-89603-191-3

  • Online ISBN: 978-1-59259-495-5

  • eBook Packages: Springer Protocols

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