RNA was extracted from cell lines as described previously, according to the manufacturer’s instructions [48 (link)]. Cells were lysed in a 6 well plate using TRIzol reagent (Invitrogen), and total RNA resuspended in RNase-free water was quantified using a Nanodrop 2000 spectrophotometer (ThermoFisher Scientific, Waltham, MA, USA). To generate cDNA for qtRT -PCR, 1 mg RNA was used with the SuperScript III First-Strand Synthesis System RT-PCR kit (Invitrogen, Waltham, MA, USA). qtRT -PCR was done as described previously, using the C1000 Touch Thermal Cycler CFX96 Real-Time System (Bio-Rad, Hercules, CA, USA). Oligonucleotide primers used for qtRT-PCR were described in [49 (link)].
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