Total RNA was isolated using the Qiagen 96-well RNeasy Prep Kit. RNA was extracted from each sample containing 1–2 × 106 cells and quality was assessed using the Agilent 2200 TapeStation system with High Sensitivity RNA ScreenTape Assay (Agilent, Santa Clara, CA). Samples with at least 5 ng/μL of RNA and an RNA integrity number score >7 were used in the library preparation following the exact protocol in the Kapa RNA HyperPrep Kit with RiboErase (Roche, Pleasanton, CA). Final libraries with a concentration greater than 4 nM and an average fragment size between 300 and 500 bases were pooled in equimolar ratio and volume to make a final sample of 4 nM. The pooled library was sequenced on a NextSeq 500/550 sequencer (Illumina, San Diego, CA) using a High Output Kit providing 150 cycles with 75 base pair paired-end reads to a minimum depth of 20 million reads per sample.