NextSeq 500 sequencing; RNA-sequencing of Aire-ILC3 cells from wild-type and Aire-deficient mice
RNA-sequencing of Aire-ILC3 cells from wild-type and Aire-deficient mice
Peripheral lymph nodes (LN) were minced and digested at 37 degrees C in RPMI (PAA) containing 0.25 mg/ml Collagenase D (Roche), 1 U/ml Dispase I (Roche), 25 mM Hepes, pH 7.2, and 120 U/ml DNase I (Roche) for 40 min, followed by addition of 5 mM EDTA and incubation for 5 min. LNs were digested with 0.5U/ml Liberase TM (Roche) and DNase I. Aire-ILC3s were sorted as Lin-, MHCII+, CD80+, IL7Ra+ cells on INFLUX sorter (BD). Total mRNA was extracted from the FACS-sorted cells using RNeasy Plus Microkit (Qiagen). The quality of the isolated RNA was controlled by Bioanalyzer 2100 electrophoresis (Agilent). RNA-seq libraries were prepared by EMBL Gene core facility in Heidelberg.
Experimental Factor: genotype
wild type genotype