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Nucleic Acids Research, 1978, Vol. 5, No. 11 4355-4374
© 1978

Articles

Most short DNA molecules isolated from 3T3 cells are not nascent

Jacek Kowalski and David T. Denhardt

Department of Biochemistry, McIntyre Medical Sciences Building, McGill University 3655 Drummond Street, Montreal, Quebec, H3G 1Y6, Canada

Received August 14, 1978. The population of short DNA molecules (less than 103 nucleotides) in 3T3 cells has been studied using in vivo and in vitro pulse labeling techniques and in vitro end-labeling. There is a large number of molecules of less than 100 nucleotides present in equal numbers in both GO and S phase cells. In S phase cells, most of these molecules are not replicating intermediates because they do not become density-labeled after a moderate period of substitution of BrdUMP, although they are detected by end-labeling in vitro This population includes the nascent Okazaki pieces that can be labeled in a short pulse with [3H]dThd or [3H]dTTP, however, these represent less than 10% of the total population. Alkaline hydrolysis of the molecules that had been end-labeled with 32P using [{gamma}32P]ATP and polynucleotide kinase did not reveal significant release of [32P] 2'(3'), 5' ribonucleoside diphosphates.


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