Dihydrofolate Reductase - Methotrexate Resistance - Normal nonmutant cultured cells are very sensitive to methotrexate (Mtx) at even about 0.1 μg/ml, which is a strong inhibitor of dihydrofolate reductase (DHFR). The Chinese hamster ovary cell line 'A29' overproduces an altered DHFR so that it is extremely resistant to Mtx. A29 DHFR encoding sequence was cloned in ρBR322 and was used to transfect Mtx-sensitive cells.

Transfected cells were selected for Mtx-resistance; some resistant lines withstood up to 40 μg/ml Mtx and contained up to 50 copies of the DHFR encoding sequence used for transfection. Clearly, DHFR gene undergoes considerable amplification during transfection.