GSM9046293: HHL5 cells treated with OA/PA for 24 h, replicate 2, m5C-MeRIP-seq; Homo sapiens; OTHER
RNA was randomly fragmented to ~200 nt using NEBNext® Magnesium RNA Fragmentation Module. Protein A/G beads were conjugated with the m5C antibody by rotating at room temperature for 1 h. The RNA fragments were then incubated with the bead-linked antibodies and rotated at 4 °C for 4 h. Following incubation, the RNA/antibody complexes were washed for several times, and the captured RNA was eluted from the complexes and purified for sequencing or qRT-PCR. RNA libraries for the IP samples were constructed with GenSeq® Low Input Whole RNA Library Prep Kit (GenSeq, Inc.) following the manufacturer's instructions. The quality of the libraries was qualified using Agilent 2100 bioanalyzer (Agilent), and the libraries were subsequently sequenced. The m5C-MeRIP-seq service and subsequent bioinformatic analysis were conducted by CloudSeq Biotech Inc. (Shanghai, China).