title: Bulk RNA-seq of Peripheral Blood Mononuclear Cells from AmaXhosa Children with Atopic Dermatitis and Healthy Controls in Rural and Urban South Africa
design: Bulk RNA-seq of Peripheral Blood Mononuclear Cells from AmaXhosa Children with Atopic Dermatitis and Healthy Controls in Rural and Urban South Africa
library_construction_protocol: Whole blood samples were collected from participating individuals (n=149) in lithium heparin tubes and PBMCs were isolated according to standard protocols. The PBMCs were separated was using Ficoll‐Paque 1.077 g/mL density gradient centrifugation. Washed PBMCs were resuspended in freezing medium (DMSO, heat‐inactivated FCS, and RPMI). Cells were then transferred to a −80 C freezer and afterwards into liquid nitrogen the following day. Total mRNA was isolated from PBMCs using the RNeasy Mini Kit (Qiagen), and RNA levels were quantified by NanoDrop (Thermo Fisher Scientific). RNA integrity was confirmed using the Agilent 2100 BioAnalyzer (Agilent). Libraries were prepared with Illumina's Truseq stranded mRNA library prep kit with polyA enrichment.