DNA Synthesis in Nucleotide Permeable Escherichia coli Cells

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Abstract
Cells made permeable to nucleotides by treatment with ether synthesize DNA from exogenous deoxynucleoside triphosphates in a semi‐conservative fashion. The reaction proceeds from sites of previous synthesis on the chromosome, and not detectably from new sites. Short chains appear as precursors of long units and have a mean sedimentation coefficient of about 10 S in alkali, like Okazaki‐pieces when synthesized in the presence of 10 to 20 μM deoxynucleoside triphosphates, and of only 5 S when synthesized in the presence of 1 μM dNTP. Under the former conditions short chains contain not more than half of the newly synthesized DNA; under the latter conditions they can contain more than half, the remainder of the newly synthesized DNA being in long units. Nascent short chains labelled after several minutes of synthesis in the presence of 5‐bromodeoxyuridine 5′‐triphosphate at high deoxynucleoside triphosphate concentrations show bimodal density distribution in an alkaline Cs2SO4 equilibrium gradient with peaks of material corresponding to the density of heavy single strands and almost light single strands, respectively. The implications of these findings are discussed. Judged from controls, repair synthesis was not a source of error in these experiments.In contrast with results reported for intact cells, ether‐treated polA1− (Kornberg‐polymerase deficient) cells join nascent short chains as efficiently as do polA+ cells.