Chromosome-mediated transfer and amplification of an altered mouse dihydrofolate reductase gene.
We have conferred methotrexate resistance on mouse 3T6 fibroblasts by chromosome-mediated transfer of an altered dihydrofolate reductase gene encoding a highly methotrexate-insensitive enzyme. The methotrexate-resistant 3T6 cell line from which the chromosomes were prepared contains multiple copies of the altered dihydrofolate reductase gene, all of which appear to reside on double-minute chromosomes. Transformants selected at 0.2 microM methotrexate contain 10-20 times more of the transferred altered gene than of the resident normal gene. The altered genes are associated with double-minute chromosomes and are permanently lost following growth of the transformants in the absence of methotrexate. Growth of the transformants in increasing concentrations of methotrexate leads to the emergence of cells which have accumulated double-minute chromosomes and which have amplified only the transferred dihydrofolate reductase gene.
D000818:Animals; D004247:DNA / Q000235:genetics; D005111:Extrachromosomal Inheritance; D005784:Gene Amplification; D007621:Karyotyping; D051379:Mice; D012641:Selection, Genetic; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics*; D014170:Transformation, Genetic
D004247:DNA; D013762:Tetrahydrofolate Dehydrogenase