Homozygous Foxn1 BAC transgenic mice on nude background
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
RNA-seq: TECs were stained using Abs against CD45 (30F11; eBioscience), EpCAM (G8.8; DSHB, University of Iowa), MHCII (M5/114.15.2; BioLegend), Ly51 (6C3; BioLegend), UEA-1 (Reactolab), Aire (5H12; eBioscience). Stained samples were sorted on a FACSAria II flow cytometer for cortical (CD45-EpCAM+MHCII+Ly51+UEA1-) and medullary (CD45-EpCAM+MHCII+Ly51-UEA1+) subpopulations and cell pelets were snap frozen in liquid nitrogen. RNA was isolated using RNeasy kit (Qiagen). ChIP-seq: TECs were enriched to 15-20% using magnetic beads (autoMACS Pro Separator, Miltenyi Biotech) and subjected to DNA crosslinking and chromatin fragmentation using truChIP Low Cell Chromatin Shearing Kit (Covaris). Chromatin was immunoprecipitated using M2 anti-FLAG antibody (F1804; Sigma) as described (Vokes et al., 2007). ATAC-seq: ~10,000 cTEC were sorted into ATAC lysis buffer and incubated with Tn5 as described in Buenrostro et al. Curr Protocol Mol Biol 2015. RNA-seq: RNA was enriched for polyA RNA, reverse transcribed and sequenced using TruSeq library prep. ChIP-seq: Library prep was carried out using TruSeq kits. ATAC-seq: Library prep was carried out as described in Buenrostro et al. Curr Protocol Mol Biol 2015