GSM7789664: TM4_DMSO_rep2; Mus musculus; RNA-Seq
RNA integrity was assessed using the RNA Nano 6000 Assay Kit of the Bioanalyzer 2100 system.Total RNA was used as input material for the RNA sample preparations. Briefly, mRNA was purified  from total  RNA using poly-T  oligo-attached magnetic beads.  Fragmentation was carried  out  using  divalent  cations  under  elevated  temperature  in  First  Strand  Synthesis Reaction Buffer(5X). First strand cDNA was synthesized using random hexamer primer and M-MuLV   Reverse   Transcriptase(RNase   H-).    Second    strand   cDNA    synthesis   was subsequently performed using DNA Polymerase I and RNase H.Remaining overhangs were converted into blunt ends via exonuclease/polymerase activities. After adenylation of 3'ends of  DNA  fragments,  Adaptor  with  hairpin  loop  structure  were  ligated  to  prepare  for hybridization. In order to select cDNA fragments of preferentially 370~420 bp in length, the library fragments were purified with AMPure XP system(Beckman Coulter, Beverly, USA). Then  PCR was performed with  Phusion  High-Fidelity  DNA polymerase,  Universal  PCR  primers and Index(X) Primer. At last, PCR products were purified (AMPure XP system) and library quality was assessed on the Agilent Bioanalyzer 2100 system. The  clustering  of the  index-coded  samples  was performed  on  a  cBot  Cluster  Generation System using TruSeq PE Cluster Kit v3-cBot-HS (Illumia) according to the manufacturer's instructions. After cluster generation, the library preparations were sequenced on an Illumina Novaseq platform and 150 bp paired-end reads were generated.