RNA Sequencing of DHOK-Treated SW620 Cells

SW620 cells were seeded in 6-cm dishes overnight and treated with 12.5 μM DHOK for 24 h in a 5% CO₂ incubator. The next day, 1 ml Trizol buffer (Beyotime, Shanghai, China) was used to lyse cells following the manufacturer’s procedure (Sun et al., 2020 (link)). The RNA sequencing (RNA-seq) experiments were undertaken by Lc-Bio Technologies Co., Ltd. (Hangzhou, China). The amount and purity of RNA was quantified by NanoDrop ND-1000 (NanoDrop, Wilmington, DE, United States). Then RNA was purified and fragmented. After that, RNA was reverse transcribed to cDNA, and next synthesized U-labeled second-stranded DNAs. An A-base was then added to the blunt ends of each strand and single- or dual-index adapters were ligated to the fragments. AMPureXP beads were used to size selection. The average insert size for the final cDNA library was 300 ± 50 bp. At last, the paired-end sequencing was performed.