We cultured HEK293T cells in RPMI medium supplemented with 10% fetal calf serum to 80% confluence in a T75. The cells were washed with 2 ml versene and incubated with 2 ml of trypsin for 3 minutes at 37 °C. We neutralized the mixture with 8 ml fresh medium. We centrifuged for 5 minutes at 2000 rcf at 4 °C and removed the supernatants. We resuspended the cells in 1 ml of QIAzol and flash-froze the mixture in liquid nitrogen.
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