For total RNA sequencing of HEK293T, RNA from SFPQ, NONO, and CTRL KD (using two different siRNAs for each condition with biological duplicates) were rRNA depleted using RiboCop rRNA Depletion Kit V1.2 (Lexogen) according to manufacturer’s protocol. Subsequent cDNA libraries were prepared using SENSE Total RNA-Seq Library Prep Kit (Lexogen) following manufacturer’s protocol.
For 3’end sequencing, cDNA libraries from HEK293T were prepared using QuantSeq 3' mRNA-Seq Library Prep Kit (Lexogen). For both methods, RNA quality was determined using the BioAnalyzer RNA nanochip (Agilent) and library concentration was quantified with KAPA Library Quant KIT RT-qPCR (Roche). Total RNAseq was done as 100nt paired-end sequencing and performed using the Illumnia platform (HiSEQ4000, BGI, Copenhagen), while for 3’end sequencing, 75nt single-end sequencing was performed at MOMA (Aarhus University Hospital) on a NextSeq500. For total RNA sequencing of HepG2 cells, library preparation and sequencing was performed at BGI (Copenhagen) using BGIseq.
For 3’end sequencing, cDNA libraries from HEK293T were prepared using QuantSeq 3' mRNA-Seq Library Prep Kit (Lexogen). For both methods, RNA quality was determined using the BioAnalyzer RNA nanochip (Agilent) and library concentration was quantified with KAPA Library Quant KIT RT-qPCR (Roche). Total RNAseq was done as 100nt paired-end sequencing and performed using the Illumnia platform (HiSEQ4000, BGI, Copenhagen), while for 3’end sequencing, 75nt single-end sequencing was performed at MOMA (Aarhus University Hospital) on a NextSeq500. For total RNA sequencing of HepG2 cells, library preparation and sequencing was performed at BGI (Copenhagen) using BGIseq.