TRANSFECTION OF A NONACTIVE SITE MUTANT MURINE DHFR CDNA (THE TRYPTOPHAN-15 MUTANT) INTO CHINESE-HAMSTER OVARY AND MOUSE MARROW PROGENITOR CELLS IMPARTS MTX RESISTANCE IN-VITRO

Chinese hamster ovary (CHO) DHFR- cells were converted to the DHFR+ (d ihydrofolate reductase) phenotype when transfected with a mammalian ex pression vector containing the murine mutant Trp15 DHFR cDNA. Transfec tion of the Trp15 DHFR cDNA into wild-type CHO cells resulted in resis tance to high levels of methotrexate (MTX) in vitro indicating that th is mutant DHFR cDNA can act as a dominant marker. Southern and Norther n blot analyses of transfected cells indicated that the transfected mu tant DHFR cDNA was integrated and expressed. Gene copy number analysis showed that the Trp15 cDNA was amplifiable in increasing concentratio ns of MTX. Transfection of murine bone marrow progenitor cells with th e Trp15 mutant DHFR cDNA resulted in MTX resistant colony forming unit s-granulocyte macrophage.