GSM1364019: TG for Nanog rep 2; Mus musculus; RNA-Seq
Total RNA was extracted combining the Trizol reagent (Life Technologies) with the RNeasy Mini kit (Qiagen). 1 µg of total RNA samples was used as provided by the user. RNA Integrity Numbers were in the range 7.2 to 8.9 when assayed on an Agilent 2100 Bioanalyzer. PolyA+ fractions were purified and randomly fragmented, converted to double stranded cDNA and processed through subsequent enzymatic treatments of end-repair, dA-tailing, and ligation to adapters as in Illumina's "TruSeq Stranded mRNA Sample Preparation Part # 15031047 Rev. D" (this kit incorporates dUTP during 2nd strand cDNA synthesis, which implies that only the cDNA strand generated during 1st strand synthesis is eventually sequenced). Adapter-ligated library was completed by 8 cycles of PCR with Illumina PE primers. The resulting purified cDNA library of template molecules of known strand origin was applied to an Illumina flow cell for cluster generation (TruSeq cluster generation kit v5) and sequenced on the Genome Analyzer IIx with SBS TruSeq v5 reagents by following manufacturer's protocols.
GEO Accession
GSM1364019