title: GSM3609817: RHP059: Peripheral blood granulocytes from heathy control; Homo sapiens; RNA-Seq
library_construction_protocol: The purified cells were lysed using mirVana total RNA isolation solution and stored at -80 °C until later RNA extraction following the manufacturer's protocol (Ambion, CA). All human RNAs were analyzed on Agilent Bioanalyzer for quality assessment with RNA Integrity Number (RIN) range from 7.1 to 10 and median RIN 9.5 (Agilent, Santa Clara, CA, USA). cDNA libraries were prepared using 300 ng of total RNA and 2ul of a 3:1000 dilution of ERCC RNA Spike in Controls (Ambion® Thermo Fisher Scientific, Waltham, MA, USA). The fragmented mRNA samples were subjected to cDNA synthesis using Illumina TruSeq® RNA sample preparation kit version 2 (Illumina, San Diego, CA, USA) according to manufacturer's protocol, except for the following modifications: 1. use of 12 PCR cycles, 2. two additional round of Agencourt Ampure XP SPRI beads (Beckman Courter) to remove >600bp double stranded cDNA. The length distribution of the cDNA libraries was monitored using DNA 1000 kits on the Agilent bioanalyzer (Agilent, Santa Clara, CA, USA). All samples were subjected to an indexed PE sequencing run of 2x51 cycles on an Illumina HiSeq 2000 (Illumina, San Diego, CA, USA) (3-4 samples/ lane). FASTQ files were mapped using STAR to the human genome build hg19. The mapped reads were then counted using featureCounts based on GENCODE V19 annotations to generate gene counts.
identifiers.namespace: GEO
identifiers.id: GSM3609817