DNA Sequencing by the Chemical Method

  • Eran Pichersky
Part of the Methods in Molecular Biology™ book series (MIMB, volume 65)

Abstract

The chemical method of sequencing DNA (1) has some advantages and some disadvantages compared with the enzymatic method (2). The major disadvantage is that it takes more time to produce the same amount of sequence. This is so for two main reasons. First, the DNA has to be end-labeled and then reisolated prior to the actual chemical sequencing reactions, a process that usually requires an additional day. Also, because more DNA is used in the reaction and because the lower specific activity of the sequenced DNA requires the use of an intensifying screen in the autoradiography, bands are not as sharp as in the enzymatic method and therefore it is difficult to obtain reliable sequence past about nucleotide 250 (unless very long gels are run).

References

  1. 1.
    Maxam, A. M. and Gilbert, W. (1980) Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 65, 499–560.PubMedCrossRefGoogle Scholar
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    Sanger, F., Nicklen, S., and Coulson, A. R. (1977) DNA sequencing with chain-terminating inhibitors. Proc. Natl. Acad. Sci USA 74, 5463–5467.PubMedCrossRefGoogle Scholar
  3. 3.
    Maniatis, T., Fritsch, E. F., and Sambrook, J. (1982) Molecular Cloning, A Laboratory Manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY.Google Scholar

Copyright information

© Humana Press Inc., Totowa, NJ 1996

Authors and Affiliations

  • Eran Pichersky
    • 1
  1. 1.Department of BiologyUniversity of MichiganAnn Arbor

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