GSM5061091: abx-04_d0_abx_MC; Homo sapiens; RNA-Seq
Bulk RNA-seq was performed on purified CD14+ monocytes after sorting. In brief, after sorting, 5,500 cells were washed, resuspended in 350 µL chilled Buffer RLT (Qiagen, 79216) supplemented with 1% beta-Mercaptoethanol (Sigma, M3148-25ML), vortexed for 1 minute, and immediately frozen at -80C. RNA was isolated using the RNeasy Micro kit (Qiagen, 74004) with on-column DNase digestion. RNA quality was assessed using an Agilent Bioanalyzer and total RNA was used as input for cDNA synthesis using the Clontech SMART-Seq v4 Ultra Low Input RNA kit (Takara Bio, 634894) according to the manufacturer's instructions. Amplified cDNA was fragmented and appended with dual-indexed bar codes using the NexteraXT DNA Library Preparation kit (Illumina, FC-131-1096). Libraries were validated by capillary electrophoresis on an Agilent 4200 TapeStation, pooled at equimolar concentrations, and sequenced on an Illumina NovaSeq6000 at 100SR, yielding 20-25 million reads per sample.