Amplified inverted duplications within and adjacent to heterologous selectable DNA.
Plasmids containing a dihydrofolate reductase (DHFR) expression unit were transfected into DHFR-deficient Chinese hamster ovary (CHO) cells. Methotrexate exposure was used to select cells with amplified DHFR sequences. Three cell lines were isolated containing amplified copies of transfected DNA that had integrated into the Chinese hamster genome. Plasmid DNA was found to co-amplify with flanking hamster sequences that were repetitive (2 cell lines) and unique (1 cell line). Fragments comprising the junctions of amplified plasmid and CHO DNA were found to exist as inverted duplications in all three cell lines. These observations provide evidence that inverted duplication occurred prior to DNA amplification, thus underscoring the importance of inverted duplication in the DNA amplification process.
Heartlein
M W
MW
Genetics Division, Children's Hospital, Boston, MA.
Latt
S A
SA
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D005784:Gene Amplification; D005796:Genes; D008727:Methotrexate / Q000494:pharmacology; D009693:Nucleic Acid Hybridization; D010957:Plasmids; D012091:Repetitive Sequences, Nucleic Acid; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics*; D014162:Transfection
D004247:DNA; D013762:Tetrahydrofolate Dehydrogenase; D008727:Methotrexate