GSM5819064: mLN WT stromal cell; Mus musculus; RNA-Seq
Single CD45‑CD24‑ cells were sorted by FACS ARIA III (BD) and collected in PBS containing 0.04 % w/v BSA at a density of 400 cells/μl. Chromium™ Controller was used for partitioning single cells into nanoliter-scale Gel Bead-In-EMulsions (GEMs) Single Cell 3' reagent kit v2 for reverse transcription, cDNA amplification and library construction (10xGenomics, Cat. #120236). The detailed protocol was provided by 10xGenomics. SimpliAmp Thermal Cycler was used for amplification and incubation steps (Applied Biosystems). Libraries were quantified by QubitTM 3.0 Fluometer (ThermoFisher) and quality checked using Fragmentanalyzer with High Sensitivity DNA kit (Agilent). Sequencing was performed in asynchronous paired-end mode (28-10-10-91 cycles) using a NovaSeq6000 sequencer (Illumina) to attain approximately 80,000 reads per single cell.