GSM3724902: Human stimluated TC rep1; Homo sapiens; RNA-Seq
Cells were lysed in RLT buffer (Qiagen Ltd., UK), tthen RNA extracted using the Qiagen's RNA Mini Kit according to the manufacturer's instructions. Briefly, RNA quality and quantity were assessed using the Bioanalyzer 2100 RNA pico kit (Agilent technologies). The input total RNA was normalized to 100 pg per sample and >9 RIN. RNA was reverse transcribed and cDNA amplified by in vitro transcription with the standard PolyA selection at the Wellcome Trust Human Genomics Centre, Oxford. Following first strand synthesis, cDNA was amplified 12 cycles by LD PCR using blocked PCR primers. Amplified cDNA was purified using AMPure XP prior to QC using the bioanalyser 2100 HS DNA kit (Agilent technologies). Library preparation of purified amplified cDNA was performed using Nextera XT library preparation (Illumina). Following QC, 100 pg of cDNA was tagmented (simultaneously fragmented with adaptors inserted) using Nextera transposons. Molecular barcodes were incorporated during 12 cycles of amplification followed by purification using AMPure XP. The libraries passed a quality checkpoint (Qubit and Bioanalyser HS DNA) prior to normalization and pooling before loading onto the HiSeq 4000 (Illumina, Oxford) for paired end sequencing.
GEO
GSM3724902