title: GSM3211573: 014-B; Human gammaherpesvirus 8; RNA-Seq
library_construction_protocol: Total RNA was isolated using the RNeasy mini kit with DNAse treatment step following manufacturer instructions. Total RNA from the 6 tumor samples analyzed by stranded sequencing were extracted using the NucleoSpin RNA kit (Machery-Nagle, Bethlehem, PA, USA). RNA was further concentrated and purified using the RNA Clean and Concentrator kit (Zymo Research, Irvine, CA, USA). Unstranded RNA-seq libraries were prepared from 300 ng of total RNA using the TruSeq RNA Sample Prep Kit v2. In addition four KS tumor libraries were prepared using the TruSeq Stranded mRNA Library Kit (Illumina) from 100 ng of total RNA.  Library size distributions were validated using an Agilent 2200 TapeStation (Agilent Technologies, Santa Clara, CA, USA).  Additional library QC, pooling of indexed libraries, and cluster optimization was performed using Life Technologies' Invitrogen Qubit® 2.0 Fluorometer (Life Technologies-Invitrogen, Carlsbad, CA, USA).  The unstranded RNA-seq libraries were pooled (5-plex) and the stranded libraries were pooled (4-plex) and each pool was clustered onto a flow cell lane.  Sequencing was performed using an Illumina HiSeq 2500 in “High Output” mode with a paired-end, 50 base reads (PE50) sequencing strategy for the unstranded libraries and non-paired end for the stranded libraries.  Image analysis and base calling was performed using Illumina's Real Time Analysis v1.18 software, followed by 'demultiplexing' of indexed reads and generation of FASTQ files, using Illumina's bcl2fastq Conversion Software v1.8.2.
attributes.tag: GEO Accession
attributes.value: GSM3211573