GSM8118124
Hela cells were scraped and resuspended in TRIZOL and were then sonicated using a Covaris m220 Ultrasonicator using default settings at 10% duty for 5 min. Then fragmented RNA was extracted using Phenol/Chloroform/isoamyl alcohol and isopropanol method. The purified RNA was kept in -20℃ fridge or directly using for library construction. The 2 μg total RNA sample was then divided into two parts for 150 mM Li+, and 150 mM K+ in Caco buffer in 60 ℃ within 5 min. Five percent of the incubated sample was collected as input control. Next, samples were incubated with 100 μM COL in 5 min after diluted in 10 mM of Li+ or K+. COL-containing samples were centrifuged for 5 min (4℃, 10000 r.p.m.) and supernatant was removed. The precipitate was resuspended with tris-HCl buffer (pH 8.8, 100 mM LiCl) and then was extracted using Qiagen RNeasy Plus Mini Kit following the manufacturer's protocol. Samples were ribo-depleted using the Ribo-off® rRNA Depletion Kit (Human/Mouse/Rat) (Vazyme). RNA library preparation was performed using the Hieff NGS® Ultima Dual-mode RNA Library Prep Kit (Yeasen) by following the manufacturer's instructions.