GSM1619508: PM19_T3; Homo sapiens; RNA-Seq
Total RNA was extracted using the mirVana™ miRNA Isolation Kit (Ambion®). 500 ng of total RNA were used to prepare the library with TruSeq RNA sample Prep Kit V2 (Illumina). Briefly, mRNA was selected from total RNA by poly-T oligo-attached magnetic beads and fragmented. Reverse transcription was done with random hexamers by SuperScript II reverse transcriptase. RNA libraries were prepared for sequencing using standard Illumina protocols.The generated libraries were submitted to quality control on High Sensitivity DNA chip from the Agilent Bioanalyzer 2100 (Agilent Technologies, Wokingham, UK) and quantified by fluorimetry using the Quant-iT™ dsDNA Assay Kit (Life Technology). A dilution at 10nM was performed for each library and pools of 12 libraries were constituted. Each pool included the libraries from 3 individuals for the 2 time points, each library being identifiable by a specific index. The resulting pools were loaded on a High Sensitivity DNA chip (Agilent) and quantified by integration. 9 pM of this solution was used for cluster generation using the TruSeq PE Cluster Kit v3 (Illumina) on a CBot instrument. A 75 bp paired-end run was performed on the Illumina HiSeq2000 platform with the TruSeq SBS kit V3 (Illumina). Sequencing was performed on two 8-laned flow cells. Each flow cell included 4 pools sequenced twice in 2 different lanes.
GEO Accession
GSM1619508