Sequencing of PCR Products — Analysis of Factor IX Genes and of Recombination Events in Immunoglobulin Genes

  • Jean-Marie Buerstedde
  • Steve S. Sommer


The polymerase chain reaction facilitates DNA sequence analysis due to the simplicity and high sensitivity of the procedure. In many cases the detection of a previously characterized sequence, restriction site or sequence polymorphism is the only information required. This information is most conveniently obtained either by determination of the size of the amplified product on an agarose gel, analysis after restriction enzyme digestion or hybridization using allele-specific oligonucleotides as probes.


Gene Conversion Carrier Testing Light Chain Gene Splice Site Junction Light Chain Locus 
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.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    Stoflet, E. S., Koeberl, D. D., Sarkar, G., and Sommer, S. S. (1988). Genomic amplification with transcript sequencing. Science 239, 491–494.PubMedCrossRefGoogle Scholar
  2. 2.
    Wong, C. Dowling, C. E., Saiki, R. K., Higuchi, R. G., Erlich, H. A., and Kazazian, H. H. (1987). Characterization of B-thalassaemia mutations using direct genomic sequencing of amplified single copy DNA. Nature 330, 384–386.PubMedCrossRefGoogle Scholar
  3. 3.
    Gyllenstein, 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. Natl. Acad. Sci. (USA) 85, 7652–7656.CrossRefGoogle Scholar
  4. 4.
    Olson, D. B., and Eckstein, F. (1989). Incomplete primer extension during in vitro DNA amplification catalyzed by Taq polymerase; exploitation for DNA sequencing. Nucleic Acids Res. 17, 9613–9620.CrossRefGoogle Scholar
  5. 5.
    Tahara, T., Kraus, J. P., and Rosenberg, L. E. (1990). Direct DNA sequencing of PCR amplified genomic DNA by the Maxam-Gilbert Method. BioTechniques 8, 366–368.PubMedGoogle Scholar
  6. 6.
    Sarkar, G., and Sommer, S. S. (1989). Access to a messenger RNA sequence or its protein product is not limited by tissue or species specificity. Science 244, 331–334.PubMedCrossRefGoogle Scholar
  7. 7.
    Sarkar, G., and Sommer, S. S. (1990). The megaprimer method of site-directed mutagenesis. BioTechniques 8, 404–407.PubMedGoogle Scholar
  8. 8.
    Koeberl, D. D., Bottema, C. D., Buerstedde, J.-M., and Sommer, S. S. (1989). Functionally important regions of the factor IX gene have a low rate of polymorphism and a high rate of mutation in the dinucleotide CpG. Am. J. Hum. Genet. 45, 448–457.PubMedGoogle Scholar
  9. 9.
    Koeberl, D. D., Bottema, C. D. K., Ketterling, R. P., Bridges, P. J., Lillicrap, D. P., and Sommer, S. S. (1990). Mutations causing hemophilia B: direct estimate of the underlying rates of spontaneous germline transitions, trans versions, and deletions in a human gene. Am. J. Hum. Genet., (in press).Google Scholar
  10. 10.
    Bottema, C. D., Koeberl, D. D., and Sommer, S. S. (1989). Direct carrier testing in 14 Families with Hemophilia B. Lancet, 526–529.Google Scholar
  11. 11.
    Tonegawa, S. (1983). Somatic generation of antibody diversity. Nature 302, 575–581.PubMedCrossRefGoogle Scholar
  12. 12.
    Reynaud, C. A., Anquez, V., Grimai, H., and Weill, J.C. (1987).A hyperconversion mechanism generates the chicken light chain preimmune repertoire. Cell 48, 379–388.PubMedCrossRefGoogle Scholar
  13. 13.
    Buerstedde, J.-M., Reynaud, C.-A., Humphries, E. H., Olson, W., Ewert, D. L., and Weill, J. C. (1990). Light chain gene conversion continues at high rate in an ALV-induced cell line. EMBO J. 9, 921–927.PubMedGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 1991

Authors and Affiliations

  • Jean-Marie Buerstedde
  • Steve S. Sommer

There are no affiliations available

Personalised recommendations