GSM7091356: Huh7 cells, Control, Day 6, Replicate 2; Homo sapiens; RNA-Seq
Cells were extracted using miRVana commercial kit, following manufacturer's instructions for total RNA isolation. Total RNA samples (250ng) were used as provided by the user. Sample RNA Integrity numbers were 8.8 on average (range 5.0-9.8) when assayed on an Agilent 2100 Bioanalyzer. Sequencing libraries were prepared with the "QuantSeq 3' mRNA-Seq Library Prep Kit (FWD) for Illumina" (Lexogen, Cat.No. 015) by following manufacturer instructions. Library generation is initiated by reverse transcription with oligodT priming, and a second strand synthesis is performed from random primers by a DNA polymerase. Primers from both steps contain Illumina-compatible sequences. Libraries are completed by PCR {This kit generates directional libraries stranded in the sense orientation (the read1 (the only read in single read format) has the sense orientation (--library-type fr-secondstrand in TopHat, --stranded=yes in HTSeq).}  cDNA libraries are purified, applied to an Illumina flow cell for cluster generation and sequenced on an Illumina instrument (see below) by following manufacturer's protocols.