title: GSM7321065: reticulocytes, HbSS samp22; Homo sapiens; RNA-Seq
library_construction_protocol: Whole blood (8-16 mL) was collected using ACD as an anticoagulatent. Plasma and platelets were isolated by gentle centrifugation at 400 g. Blood was further layered on ficoll-histopaque (density 1.077 g/mL) for separation of mononuclear cells and lymphocytes. The lower layer of packed erythrocytes were transferred to a clean tube and preferentially lysed with 4 volumes of 0.144 M ammonium chloride, 1 mM ammonium bicarbonate solution. Granulocytes were separated by centrifugation and the supernatant containing reticulocyte RNA was precipitated by acid titration to pH 5.1. Precipitated RNA was preserved in Trizol and stored at -80ºC Illumina TruSeq Stranded Total RNA Sample Preparation Kit with Ribo-Zero Human/Mouse/Rat. Strand-specific libraries were constructed for 100 or 125 bp paired-end sequencing to 30-45 million read pairs per sample