Cloning and characterization of DNA sequences amplified in multidrug-resistant Djungarian hamster and mouse cells.
Five recombinant phages containing different parts of genomic regions amplified in multidrug-resistant (MDR) cells have been isolated from genomic libraries of colchicine- and actinomycin D-resistant Djungarian hamster cells. Fragments of these clones together with a part of Chinese hamster mdr gene (plasmid pDR4.7 a gift of Dr. I. Roninson) were used as hybridization probes to study composition and variations of amplified DNA in a large number of MDR cell lines. Two of the six probes used (pC52 and pDR4.7) showed DNA amplification in a large number of MDR cell lines tested (commonly amplified clones) regardless of their origin (Djungarian hamster or mouse), type of selective agent used (colchicine, actinomycin D, or anthracyclines), and mode of selection (in vitro or in vivo). These clones hybridized with two different RNA transcripts (pDR4.7, 5kb; pC52, 3.5 kb) that were overproduced in MDR cells. Degrees of amplification of both commonly amplified sequences correlated with levels of resistance in all but one of the cell lines. Other cloned sequences (sporadically amplified clones) were amplified to different extents (but never greater than the commonly amplified sequences) in some of the Djungarian hamster MDR cell lines. Such differential amplification is not the result of heterogeneity of cell population since 20 cell clones tested showed identical ratios of amplification of different amplified sequences. Sporadically amplified sequences usually coamplified with the commonly amplified ones at first steps of selection, but then they would cease to amplify and, at the later stages of selection, they could even be completely deamplified. It seems that disappearance of unnecessary parts of amplicons is a regular process accompanying stepwise gene amplification.
Gudkov
A V
AV
Laboratory of Oncovirology, Cancer Research Center, Moscow, U.S.S.R.
Chernova
O B
OB
Kazarov
A R
AR
Kopnin
B P
BP
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D006224:Cricetinae; D004247:DNA / Q000235:genetics*; D004262:DNA Restriction Enzymes; D004351:Drug Resistance / Q000235:genetics*; D005784:Gene Amplification; D051379:Mice
D004247:DNA; D004262:DNA Restriction Enzymes