RNA-seq of MYC-Low and MYC-High FC-MYC Cells Following SUMOylation Inhibition with SAEi ML-792
RNA-seq of MYC-Low and MYC-High FC-MYC Cells Following SUMOylation Inhibition with SAEi ML-792
Human colon carcinoma cell line RKO (ATCC; Cat#CRL-2577) was used. Both endogenous MYC alleles were mutated using CRISPR/Cas9-mediated gene editing. Then, Flp-mediated recombination was used to insert a tetracycline-responsive MYC transgene into a pre-existing FRT site, thus generating CRISPR-Flp-MYC cells RKO cells were cultured in Dulbecco's Modified Eagle Medium (DMEM). RKO Flp-InTM T-RexTM FC7-MYC cells were cultured in DMEM with 100 ng/ml tetracycline, 8 μg/ml blasticidin and 400 μg/ml hygromycin B (Sigma Aldrich), as described previously (Littler et al., 2019).  RKO Flp-InTM T-RexTM FC-MYC cells were seeded into T75 flasks at 300,000 (MYC-high) or 400,000 (MYC-low) cells/flask with either DMSO or 25 nM SAEi ± 500 ng/mL tetracycline. After 24 hours, cells were scraped into the media, centrifuged and resuspended in PBS, pelleted and stored at -80°C before RNA extraction RNA extraction was performed using RNeasy Micro Kit (Qiagen) following manufacturer's instructions. RNA was quantified using a NanoDrop One (Thermo Scientific) and 1 μg stored at -80°C Libraries were generated using the TruSeq Stranded mRNA assay (Illumina, Inc.) following manufacturer's instructions. Adaptor indices were used to allow multiplexing of pooled libraries before cluster generation using a cBot instrument (Illumina, Inc.).