GSM7949406: NC; Mus musculus; OTHER
Cell supernatants were centrifuged at 10,000 ×g for 45 min at 4 °C to remove the larger vesicles. The supernatants were extracted and filtered through a 0.45 µm filter. Then, the samples were centrifuged again at 10,000 ×g and at 4 °C for 70 min and resuspended in PBS. The m7G-tRNA MeRIP-Seq service was provided by CloudSeq Inc. (Shanghai, China). Briefly, total RNA was purified for small RNA fraction ( <200nt) with the MirVana Isolation Kit (ThermoFisher). The anti-m7G MeRIP was performed on the small RNA with GenSeq® m7G MeRIP Kit (GenSeq, Inc.) by following the manufacturer's instructions except omit the fragmentation procedure. Then, the MeRIP/Input small RNAs were demethylated with AlkB enzyme mix for 100 min at 37°C. The demethylated small RNA was used for library construction with GenSeq® Small RNA Library Prep Kit (GenSeq, Inc.) by following the manufacturer's instructions. All libraries were size-selected for tRNA fraction before sequencing and then sequenced in a NovaSeq platform (Illumina, Inc., San Diego, CA, USA).