title: GSM5213214: Spleen CD32- T4 cells from CM #2; Macaca fascicularis; RNA-Seq
library_construction_protocol: RNA was isolated from the sorted CD4 populations cells using the RNeasy® Mini Kit (205113, Qiagen) RNA integrity was verified with the Agilent Bioanalyser. DNase-treated RNA was treated for library preparation using the Truseq Stranded mRNA Sample Preparation Kit (Illumina, San Diego, CA), according to manufacturer's instructions. An initial poly(A) RNA isolation step (included in the Illumina protocol) is performed on 10 ng of total RNA to keep only the polyadenylated RNA fraction and remove the ribosomal RNA. A step of fragmentation is then performed on the enriched fraction, by divalent ions at high temperature. The fragmented RNA samples were randomly primed for reverse transcription followed by second-strand synthesis to create double-stranded cDNA fragments. No end repair step was necessary. An adenine was added to the 3'-end and specific Illumina adapters were ligated. Ligation products were submitted to PCR amplification. The obtained oriented libraries were controlled by Bioanalyzer DNA1000 Chips (Agilent, # 5067-1504) and quantified by spectrofluorimetry (Quant-iT™ High-Sensitivity DNA Assay Kit, #Q33120, Invitrogen). Sequencing was performed on the Illumina Hiseq2500 platform to generate single-end 100 bp reads bearing strand specificity.
attributes.tag: GEO Accession
attributes.value: GSM5213214