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DNA Sequencing by the Chemical Method

  • Eran Pichersky
Protocol
Part of the Springer Protocols Handbooks book series (SPH)

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).

Keywords

Reaction Tube Pasteur Pipet Stop Solution Heat Block Dimethyl Sulfate 
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.

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
  2. 2.
    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. (eds.) (1982) Molecular Cloning, A Laboratory Manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.Google Scholar

Copyright information

© Humana Press Inc., Totowa, NJ 2000

Authors and Affiliations

  • Eran Pichersky
    • 1
  1. 1.Department of Biology, Natural Sciences BuildingUniversity of MichiganAnn Arbor

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