GSM9307210: hESC, METTL3 KD, rep2; Homo sapiens; RNA-Seq
Total RNA from the samples was extracted using TRIzol, and RNA was purified according to the MEGA clear transcription purification kit. RNA was incubated with fragmentation buffer for 4 min at 94°C and glyoxal solution and DMSO were added, and the mixture was incubated for 30 min at 50°C in a preheated thermal cycler. Minutes, and at the end of the incubation, the tubes were brought to room temperature, and then saturated H3BO3 solution was added. The incubation was continued at 50 °C for 30 min. Then add to the pre-configured deamination buffer (750 mM NaNO2, 40 mM MES (pH 6.0), and 10 μl of glyoxal solution) for a total of 8 h of incubation. Finally, RNA was purified by ethanol precipitation, and sequencing libraries were constructed by GenSeq® Low Input RNA Library Prep Kit following the process of the instructions provided by the manufacturer. The constructed sequencing libraries were subjected to quality control and quantification by BioAnalyzer 2100 system, followed by 150 bp double-end sequencing.