title: GSM8001221: HSV-IP1; Homo sapiens; RIP-Seq
library_construction_protocol: RIP-Seq: The cells were collected into 1 ml of a polysome lysis buffer containing 10 mM HEPES (pH7.4), 100 mM KCl, 0.5% Nonidet P-40, 0.1% SDS, 0.5 mM DTT, and a cocktail of Roche EDTA-free protease inhibitors. The lysates were cleaned by centrifugation and used for immunoprecipitation by incubating at 4 °C with the antibody pre-bound to BeyoMag Protein A/G magnetic beads (P2108, Beyotime). The beads were washed with a buffer (50 mM Tris-HCl, pH 7.4, 150 mM NaCl, 1 mM MgCl2, 1 mM DTT) for 5 times and then resuspended for proteinase K treatment (55 °C for 30 min). The supernatants were collected, treated with DNase I at 37 °C for 10 min to remove potential DNA contamination, and then cleaned by phenol-chloroform extraction. The RNA was ethanol precipitated and reverse transcribed into cDNA using HiScript® III SuperMix (Vazyme) and usedfor library construction. The RNA-seq library preparation was followed by the instructions of genergy bio-technology CO., Ltd. RNA-Seq: Total cellular RNA was extracted using the RNA Rapid Extraction Kit (Vazyme, Nanjing, China) and total tissue RNA was extracted using Trizol (Vazyme), according to the reagent instructions.