GSM6705146: CF, sorted lung IMs, after chronic LPS, rep1; Mus musculus; RNA-Seq
Lungs of treated WT and CF mice were perfused to remove circulating blood cells and single cell suspension from lung tissues were isolated using a lung dissociation kit (Miltenyi). Cells were stained and FACS-sorted, and RNA was extracted using miRNEasy Micro kit (Qiagen, MD). Markers used to sort monocytes/macrophages are: cMons: CD45+CD11b+MHC-II-Ly6C+; IMs: CD45+CD11b+MHC-II+; moAMs: CD45+CD11c+SiglecFdim; trAMs: CD45+CD11c+SiglecF+ The RNA integrity (RIN) and quantity was determined on the Agilent 2100 Bioanalyzer (Agilent, CA) with Agilent RNA 6000 Pico Kit (Agilent, CA). Only samples with RIN ≥8.0 were used for library preparation. Reverse transcription and cDNA preamplification were performed using the SMART-Seq v4 Ultra Low Input RNA Kit for Sequencing (Takara Bio USA, MI), and sequencing libraries were prepared using Nextera XT DNA Sample Preparation kit (Illumina, CA, USA), according to manufacturers' instructions. Libraries were pooled and sequenced on Illumina HiSeq4000 (Illumina, CA, USA) using paired-end sequencing for 100 cycles.