Cells were collected with PBS at 4 ° C, centrifuged at 4 ° C, 1000 g, and 5 min and the supernatant was discarded and placed in liquid nitrogen. The enriched RNA was fragmented into short fragments using Fragmentation Buffer. Then a cDNA strand was synthesized using six-base random primers based on the fragment mRNA template, and then a two-strand cDNA was synthesized by adding buffer, dNTPs and DNA polymerase I. The eluted and purified double-stranded cDNA was then treated with terminal repair, base A and sequencing splice. The target size fragments were recovered by magnetic beads and amplified by PCR. Qubit 2.0 was used to quantify the libraries, and then different libraries were sequenced on Illumina sequencers according to the requirements of effective concentration and target on-machine data volume.