GSM2125477: DF508-1; Mus musculus; RNA-Seq
When the cells become confluent in 10cm dish at Passage 4, cells were collected after washing with PBS. RNA was harvested using Trizol reagent. The total RNA samples are first treated with DNase I to degrade any possible  DNA contamination. Then the mRNA is enriched by using the oligo(dT) magnetic beads. Mixed with the fragmentation buffer, the mRNA is  fragmented into short fragments. Then the first strand of cDNA is synthesized by using random hexamer-primer. Buffer, dNTPs, RNase H and  DNA polymerase I are added to synthesize the second strand. The double strand cDNA is purified with magnetic beads. End reparation and 3’-  end single nucleotide A (adenine) addition is then performed. Finally, sequencing adaptors are ligated to the fragments. The fragments are  enriched by PCR amplification. During the QC step, Agilent 2100 Bioanaylzer and ABI StepOnePlus Real-Time PCR System are used to  qualify and quantify of the sample library. The library products are ready for sequencing via Illumina HiSeqTM 2000.
GEO Accession
GSM2125477