GSM7988411: mac_disease NFS bio rep 3; Homo sapiens; RNA-Seq
Up to 1.2 x106 CD14+ monocytes or monocyte-derived macrophages from patients and healthy controls were carefully washed, harvested and lysed in 700 ul Qiazol (Qiagen) and stored at -80°C until RNA isolation. CD14+ cells were lysed on the day of sample collection, whereas macrophages were harvested after 3 days of differentiation with GM-CSF. RNA was isolated from the aforementioned samples using the miRNeasy Kit (Qiagen) and concentration and purity assessed using a Nanodrop 2000c Spectrophotometer (Thermo Scientific). RNA quality of the samples was further assessed on the Agilent Bioanalyzer RNA 6000 Nano Electropherogram, and library preparation performed on samples with RIN quality between 8-10. 100ng of total RNA was used for library preparation using the NEBNext Ultra II directional RNA library preparation kit with poly-A tail enrichment. stranded mRNA 75 bp single read RNA sequencing was performed on the Illumina NextSeq 500 sequencer.