title: RNA-Seq of TE-9 cell line
design: After the RNA sample is qualified, use magnetic beads with Oligo (dT) to enrich eukaryotic mRNA (if it is a prokaryote, remove rRNA through the kit to enrich mRNA). Subsequently, fragmentation buffer is added to break the mRNA into short fragments. Using mRNA as a template, one-strand cDNA is synthesized with six-base random primers (random hexamers), and then buffer, dNTPs, and DNA polymerase I and RNase H are added to synthesize two-strand cDNA. Then use AMPure XP beads to purify double-stranded cDNA. The purified double-stranded cDNA is first repaired, A-tailed and connected to the sequencing adapter, and then AMPure XP beads are used for fragment size selection. Finally, perform PCR amplification, and use AMPure XP beads to purify the PCR products to obtain the final library. After the library is constructed, use Qubit 2.0 for preliminary quantification, dilute the library, and then use Agilent 2100 to detect the size of the insert in the library. After the insert meets expectations, use the Q-PCR method to accurately quantify the effective concentration of the library. Ensure the quality of the library. After the library is qualified, the different libraries are pooled to flowcell according to the effective concentration and target data volume requirements. After the cBOT is clustered, the Illumina high-throughput sequencing platform NovaSeq 6000 is used for sequencing.