GSM8022760: CCB105, YF-17D vaccination, day 28, rep 1; Macaca fascicularis; RNA-Seq
Whole blood from cynomolgus macaques was collected in PAXgene tubes (BD762165 –Ozyme), and stored according to the manufacturer's protocol. RNA was extracted with the PAXgene 96 Blood RNA kit (Qiagen-762331) according to manufacturer's instructions on Tecan Evo150 workstation. The RNA quantity, purity and quality were checked on the Labchip (Caliper) and the Nanodrop Spectrophotometer (Thermo Scientific). Extracted RNA samples with an A260/A280 ratio ~2, a RIN>8 and a minimal concentration of 70ng/µL of RNA were processed using the GLOBINclear™-Human kit (ThermoFisher - AM1980) for the depletion of the alpha and beta globin mRNA according to manufacturer's instructions. The quantity, purity and quality of the RNA depleted of globin mRNA was reassessed as already described before library preparation for sequencing. Sequencing libraries were prepared with the TruSeq mRNA stranded kit (Illumina - 20162122/20146602) using 300ng of RNA according to manufacturer's instructions. The quality, length distribution and quantity of the resulting libraries were analyzed with the Qubit 3.0 Fluorometer (Life Technologies) and the Labchip (Caliper). Libraries were pooled at equimolar amounts and sequenced by single reads of 75bp on high-output flowcell (Illumina – FC-404-2005) using the Nextseq500 sequencer (Illumina) in a batch of 10 samples per flowcell with the specification to achieve 30 million reads per sample. On run completion, libraries were demultiplexed, adapters trimmed and FASTQ files generated.