Normal primary cultures of human thyroid follicular cells were transfected with a plasmid containing the SV40 early region large T encoding gene from mutant tsA58. Tight temperature dependence for growth is observed: Nthy-ts1 cells grow at 33°C, but not at the restrictive temperature of 40.5°C. This SV40 mutant allows the study of oncogene action free of any interfering effects of SV40 large T antigen.
Split sub-confluent cultures (70-80%) 1:3 to 1:6 i.e. seeding at 2-4x10,000 cells/cm2 using 0.25% trypsin or trypsin/EDTA; 5% CO2; 33°C.
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