Nucleotide sequence analysis of DNA. XXIV. Synchronous digestion of SV40 DNA by exonuclease III

Authors:
  • RWRay Wu
    Ray Wu
  • GRGeorge Ruben
    George Ruben
  • BSBenjamin Siegel
    Benjamin Siegel
  • EJErnest Jay
    Ernest Jay
  • PSPaul Spielman
    Paul Spielman
  • CTChen-Pei D. Tu
    Chen-Pei D. Tu
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Abstract
We have established an optimal condition for the synchronous digestion of SV40 DNA with Escherichia coli exonuclease III. Electron microscopy and polyacrylamide gel electrophoresis were used to obtain accurate measurements on the lengths of DNA before and after exonuclease III digestion. Based on this finding, a new method for determining the sequence of long duplex DNA can be realized. It involves (a) the synchronous digestion of the DNA from the 3' ends with exonuclease III, followed by (b) repair synthesis with labeled nucleotides and DNA polymerase, and (c) sequence analysis of the repaired DNA.