Optimal conditions for supercoil DNA sequencing with the Escherichia coli DNA polymerase I large fragment

Gene Anal Tech. 1988 Mar-Apr;5(2):32-9. doi: 10.1016/0735-0651(88)90024-6.


Sequencing ladders produced from supercoiled DNA templates with the Escherichia coli DNA polymerase Klenow fragment are often unreadable because of a high background and misincorporated nucleotides. This study showed that contaminating RNA molecules can interfere with template:primer hybridization. Procedures are provided for the purification of template DNA and stringent conditions for primer-template hybridization that overcome these problems.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • DNA Polymerase I / metabolism*
  • DNA, Superhelical / genetics*
  • Electrophoresis, Polyacrylamide Gel
  • Escherichia coli / enzymology*
  • Plasmids
  • Templates, Genetic


  • DNA, Superhelical
  • DNA Polymerase I