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Molecular Structure and Evolution of Double-Minute Chromosomes in Methotrexate-Resistant Cultured Mouse Cells by Peter J. Hahn; Barbara Nevaldine; John A. Longo is a Biochemistry, Genetics and Molecular Biology article available to read on EtoBox.

What is Molecular Structure and Evolution of Double-Minute Chromosomes in Methotrexate-Resistant Cultured Mouse Cells about?

To determine whether microscopically visible double-minute chromosomes (DMs) are derived from submicroscopic precursors, we monitored the amplification of the dihydrofolate reductase (DHFR) gene in 10 independent isolates of methotrexate (MTX)-resistant mouse cells. At every other doubling in MTX concentration, the cells were examined both microscopically, to detect the presence of microscopically visible DMs, and by pulsed-field gel electrophoresis and hybridization to a DHFR-specific probe, to detect submicroscopic DMs. One of the cloned MTX-resistant isolates was examined in detail and was shown to originally contain amplified DHFR genes on circular DMs measuring 1 and 3 Mb in size; additionally, metaphase chromosome preparations from this cloned isolate were examined and were shown to contain microscopically visible DMs too large to enter a pulsed-field gel. During stepwise selection for increasing levels of MTX, the smaller DMs (not microscopically visible) were shown to be preferentially amplified, whereas the larger (microscopically visible) ones decreased in relative numbers. Rare-cutting NotI digestion patterns of total genomic DNA that includes the DMs containing the DHFR

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Author

Peter J. Hahn; Barbara Nevaldine; John A. Longo

Publisher

Informa UK Limited

Published

1992

Language

EN

Field

Biochemistry, Genetics and Molecular Biology (Life Sciences)