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Nucleic Acids Research, 1978, Vol. 5, No. 2 331-347
© 1978

Articles

Production of DNA bifilarly substituted with bromodeoxyuridine in the first round of synthesis: branch migration during isolation of cellular DNA

K. Tatsumi and B. Strauss

Department of Microbiology, The University of Chicago 920 East 58th Street, Chicago, IL 60637, USA

Received October 21, 1977. Incubation of human lymphoid cells with bromodeoxyuridine (BrdUrd) for short periods produces three classes of DNA containing analog: DNAHL (hybrid DNA, density{cong}1.75 g/cm3), DNAint (intermediate density DNA, density{cong}1.71 g/cm3), and DNAHH (DNA with both strands containing analog, density{cong}1.80 g/cm3). Preparations of DNAint yield DNAHH after extensive shearing and/or treatment with single strand specific endonuclease. Cross-linking of pulse-labeled (Brdurd+3HdT) DNA in cells by treatment with trioxsalen and near UV light before lysis prevents the appearance of DNAHH. Cross-linking, after lysis has little effect. A large fraction of DNAHH is obtained after incubation of cells with caffeine. Extraction of DNA at high salt concentration or cross-linking with trioxsalen and near UV light drastically reduced the amount of DNAHH obtained from caffeine-treated cells. We conclude that most DNAHH arises from in vitro branch migration in isolated DNA growing points.


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