Illumina NovaSeq 6000 paired end sequencing; Comparison of human plasmacytoid dendritic cells and monocyte-derived dendritic cells
Comparison of human plasmacytoid dendritic cells and monocyte-derived dendritic cells
Immediately after thawing, PBMCs were stained with antibody cocktails and pDCs were sorted by FACS according to the CD123 (IL-3R) and BDCA-2 (CD303) expressions. As an internal control, mDCs were isolated from the same PBMC donors and simultaneously sorted by FACS according to the CD11c expression.  RNA was extracted from the 1 x106 cells by QIAzol Lysis Reagent (QIAGEN). Concentrations of nucleic acids were measured by nanodrop (all experiments) and/or Qubit HS RNA-kit (only for RNA-seq). RNA quantity and quality were measured by Agilent 2100 Bioanalyzer (Agilent technologies). RNA integration values (RIN) >8 were considered to proceed to the further analysis. RNA-seq libraries were generated using Illumina TruSeq Stranded Total RNA kits. The resultant cDNA libraries were quantified via Qubit and assessed for quality via Bioanalyzer.