title: GSM9499699: engineered neural tissues (ENT), EF, Day7, rep2; Mus musculus; RNA-Seq
library_construction_protocol: Purity, concentration and integrity of RNA sample were examined by NanoDrop, Qubit 2.0, Agilent 2100, etc. Only RNA with good quality could move on to following procedures. Qualified RNA was isolated by Oligo(dT)-attached magnetic beads, then randomly fragmented in fragmentation buffer. First-strand cDNA was synthesized with fragmented mRNA as template and random hexamers as primers, followed by second-strand synthesis with addition of PCR buffer, dNTPs, RNase H and DNA polymerase I. Purification of cDNA was processed with AMPure XP beads. Double-strand cDNA was subjected to end repair. Adenosine was added to the end and ligated to adapters.  AMPure XP beads were applied here to select fragments within size range of 300-400 bp. cDNA library was obtained by certain rounds of PCR on cDNA fragments.