Highly efficient copying of single-stranded DNA by eukaryotic cell chromatin

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Abstract
Chromatin prepared from S phase hepatoma tissue culture (HTC) cell incorporates in vitro about 11-14 pmoles [3H]dTMP into DNA in 30 min. Single-stranded DNA added to this chromatin stimulates DNA synthesis more than 40-fold whereas activated DNA enhances it about 60-fold. By contrast, stimulation of DNA synthesis by activated DNA in a crude nuclear extract exceeds the stimulation exerted by denatured DNA by a factor of 7. Stimulation of DNA synthesis by denatured DNA is not due to stabilization of either the chromatin or the product of the endogenous reaction. On the other hand, we find that poly(dC) and poly (dT) enhance DNA synthesis by serving as templates which are copied by chromatin in a true complementary fashion. It seems therefore, that eukaryotic cell chromatin is able to copy single-stranded DNA at a high efficiency. Chromatin of G1 arrested cell copies exogenous templates at a considerably reduced rate. The enzyme responsible for the copying of denatured DNA is tentatively identified as DNA polymerase alpha on the basis of its sensitivity to sulfhydril group blocking, its requirements for ions and failure to copy the ribo strand of oligo(dT) poly(A).