1h-2 (CRX2129092)
Advanced molecular biology equipment is used to detect the purity, concentration and integrity of RNA samples to ensure that qualified samples are used for transcriptome sequencing. After the sample is qualified, the library is constructed. The main process is as follows: enrich eukaryotic mRNA with magnetic beads with Oligo (dT); add Fragmentation Buffer to randomly interrupt the mRNA; use mRNA as a template, use six-base random primers (random hexamers) to synthesize the first cDNA chain, and then add buffer, dNTPs, RNase H and DNA polymerase I to synthesize the second cDNA chain, and use AMPure XP beads to purify the cDNA; the purified double-stranded cDNA is then end-repaired, A-tailed and connected to the sequencing adapter, and then AMPure XP beads are used for fragment size selection; finally, the cDNA library is enriched by PCR. After the library is constructed, Qubit2.0 and Agilent 2100 are used to detect the concentration and insert size of the library, and the effective concentration of the library is accurately quantified using the Q-PCR method to ensure the quality of the library. Sequencing on the machine