GSM5243323: iNKT1 CD4neg-1; Mus musculus; RNA-Seq
Bulk RNA-seq of iNKT sublineages: RNA was extracted from freshly FACS-sorted iNKT sublineages using the RNeasy mini kit (Qiagen) after pelleting 10k-50k cells, ATAC-seq: Lysis was performed in 100 μl cold buffer (10 mM Tris-HCl, pH 7.4, 10 mM NaCl, 3 mM MgCl2 + 0.1% IGEPAL CA-630 + 0.1% Tween 20), and transposition was performed in 50 μl buffer containing 25 μL 2× TD buffer (Illumina #FC-121-1030), 2.5 μL Tn5 transposase (Illumina #FC-121-1030), 17.5 μL nuclease free H2O, and 5µl 1% digitonin. Bulk RNA-seq of conventional T cells after HCT: RNA was extracted using the TRIzol RNA isolation method (ThermoFisher Scientific) combined with the RNeasy MinElute Cleanup (Qiagen). Single-cell libraries were generated using the Chromium Controller Single-Cell Instrument and Chromium Single Cell 3′ Library & Gel Bead Kit v2 (10x Genomics). Bulk cDNA libraries of sorted iNKT sublineages were prepared using the Truseq stranded mRNA kit (Illumina) and pooled for sequencing on the Illumina HiSeq 4000 platform (75 bp, paired-end). ATAC seq was performed as previously described (Buenrostro et al., Nat. Methods 2013), following the reommended library preparation protocol and sequencing on and Illumina HiSeq 4000. Bulk RNA-seq of conventional T cells after HCT: Full-length cDNA was generated using the Clontech SMARTer v4 kit (Takara Bio USA, Inc., Mountain View, CA) prior to library generation with the Nextera XT DNA Library Prep kit (Illumina, Inc., San Diego, CA). Libraries were pooled for sequencing on the Illumina HiSeq 4000 platform (75 bp, paired-end). RNA-seq, ATAC-seq
GEO Accession
GSM5243323