Illumina HiSeq 4000 paired end sequencing; A conserved population of MHC II-restricted, innate-like, commensal-reactive T cells in the gut of humans and mice
A conserved population of MHC II-restricted, innate-like, commensal-reactive T cells in the gut of humans and mice
Colonic lamina propria lymphocytes were isolated by EDTA treatment, collagenase digestion, and 3 layer percol per standard Powrie lab protocols.  Cells were stained with antibody cocktail (see lab notebook 2018 27 and 29/07/2018) and sorted into CD127+ DN T cells, MAIT, NKT, and CD127+CD4 and Cbir DN populations. Cells were sorted into 300ul of RLT and frozen until RNA extration. Qiagen RNAeasy Micro Kit with DNAse treatment Smart-Seq2 following manufacturers instructions followed by our own Nextera indexing primers during amplification, dual  indexing tags are based on this paper (DOI: 10.1186/1472-6750-13-104). Amplified libraries were analysed for size distribution using the Agilent Tapestation 2200 D1000. Libraries were quantified using Picogreen and relative volumes were pooled accordingly.