scRNAseq_musmusculus_cell_line_mTEC3.10_WT_adh_thymocytes_36h
In vitro Aire WT mTECs:In this project, we co-culture murine Aire wild-type mTEC cells together with thymocytes (mTEC 3.10 cell line plus fresh SP TCD4+ thymocytes) for 36 hours. Co-culture showed >90% viability. Cells were processed following the 10X Genomics protocol to prepare a single-cell library using Chromium equipment (10X Genomics). Next we used the Illumina protocol: The single cells were isolated and lysed, the mRNAs were purified and primed with a poly(T) primer for reverse transcription. Unreactive primers were removed by exonuclease I digestion. Poly(A) tails were added to the first strand cDNA at the 3' end and annealed to poly(T) primers for second-strand cDNA generation. The cDNAs were PCR-amplified, sheared, and prepared into sequencing library. The library was sequenced in an Illumina NovaSeq 6000 sequencer.