GSM8816831: RAW264.7 cell, medium+10nM Dox, rep5; Mus musculus; RNA-Seq
cells in each well were lyzed by 0.5ml TRNzol Universal buffer (TIANGEN Biochemical Technology (Beijing), DP424) on a slowly swinging rocker at 4℃ for 15min, and then collected into a cryogenic storage tube and stored at -80℃. The RNA was extracted by chloroform (Sigma-Aldrich, CX1060) The mRNA in each sample was enriched by Oligo (dT) magnetic beads and fragmented by the metal ion solution. Then reverse transcription was done using the M-MuLV system and random oligonucleotide primers, and the sticky ends of end-repaired, double-strand cDNA with poly-A tail was catalyzed by T4 DNA ligase with short adapters.AMPure XP beads were used for post-ligation purification, followed by size selection of cDNA fragments around 300-350bp. Polymerase chain reaction (PCR) was conducted to incorporate sample indices, and the PCR products were purified with AMPure XP beads to obtain the final library. The library was quantified, diluted, and analyzed for fragment distribution with the Agilent 2100 DNA 1000 kit. Quality control was confirmed, and qRT-PCR was used for precise quantification of the library's effective concentration to ensure quality.