Sites of termination of in vitro DNA synthesis on ultraviolet- and N-acetylaminofluorene-treated phi X174 templates by prokaryotic and eukaryotic DNA polymerases.

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Abstract
In vitro DNA synthesis on a .vphi.X174 template primed with a restriction fragment and catalyzed by the Escherichia coli DNA polymerase I large (Klenow) fragment (pol I) terminates at the nucleotide preceding a site that has been altered by UV irradiation or treatment with N-acetylaminofluorene. Termination on UV-irradiated templates is similar when synthesis is catalyzed by E. coli DNA polymerase III holoenzyme (pol III), phage T4 DNA polymerase, a polymerase .alpha. from human lymphoma cells or avian myeloblastosis virus reverse transcriptase. 3'' .fwdarw. 5'' Exonuclease activity cannot be detected in the reverse transcriptase and DNA polymerase .alpha. preparations. On N-acetylaminofluorene templates, pol I, pol III and T4 polymerase reactions terminate immediately preceding the lesion; reverse transcriptase-catalyzed reactions and, at some positions in the sequence, polymerase .alpha.-catalyzed reactions terminate at the site of the lesion. Substitution of Mn2+ for Mg2+ changes the pattern of pol I-catalyzed termination sites. Termination evidently is a complicated process that does not depend exclusively on the 3'' .fwdarw. 5'' exonuclease activity associated with many polymerases.