Early appearance and long-term persistence of the submicroscopic extrachromosomal elements (amplisomes) containing the amplified DHFR genes in human cell lines

Submicroscopic extrachromosomal elements (amplisomes) containing amplified dihydrofolate reductase (DHFR) genes have been investigated in a methotrexate-resistance derivative of the human cell line HeLa BU25, 10B3, by field-inversion gel electrophoresis. The amount and kinetics of formation of these elements have been correlated with the level and time course of overall DHFR gene amplification. The amplisomes account for the great majority and possibly the totality of the amplified DHFR genes in 10B3 cells. They appear very early during the development of methotrexate resistance and increased in parallel with the amplified genes. These observations suggest that these elements are involved in an early event, possibly the first event, of gene amplification in this system. Amplisomes tend to be lost from 10B3 cells in the absence of selective pressure, although much more slowly than expected from simple dilution of nonreplicating elements. Surprisingly, under selective pressure, these elements have shown no tendency to become integrated into chromosomes or to generate minute chromosomes over a period of almost 1 year, in contrast to what has been described in other systems