GSM9075525: Exp3 Lane 1; Mus musculus; RNA-Seq
Experiment 1: Spleens were obtained from 7-8 week-old EpoR-loxp/loxp or EpoR-loxp/loxp-Xcr1-Cre mice after TLI/ATS treatment or from untreated controls. Samples were dissociated and subjected to MACS negative enrichment with Pan Dendritic Cell Isolation Kit and were tagged by genotpye using TotalSeq-A hashtag antibodies. Lin(CD3e, NK1.1, CD19, B200)-SiglecH-PDCA-1-F4/80-CD11c+MHCII+XCR1+SIRPa- cDC1s were FACS isolated and used for 10x Genomics scRNA-seq. Experiment 2: Spleens and LNs were obtained from 7-8 week-old EpoR-TdTomato-Cre mice after TLI/ATS treatment. Samples were dissociated and subjected to MACS negative enrichment with Pan Dendritic Cell Isolation Kit, split into 2 aliquots per tissue and MULTI-seq barcoded. cDC1s were then FACS isolated according to EpoR expression status (tracked with MULTI-seq barcode), pooled by EpoR status, and used for 10x Genomics scRNA-seq. Experiment 3: LNs were obtained from 7-8 week-old EpoR-loxp/loxp or EpoR-loxp/loxp-Xcr1-Cre mice. Samples were dissociated and subjected to MACS negative enrichment with Pan Dendritic Cell Isolation Kit and were tagged by genotpye using TotalSeq-A hashtag antibodies. Lin(CD3e, NK1.1, CD19, B200)-SiglecH-PDCA-1-F4/80-CD11c+MHCII+XCR1+SIRPa- cDC1s were FACS isolated and used for 10x Genomics scRNA-seq. Libraries were prepared according to manufacturers instructions (10x Genomics V3.1; BioLegend Total-Seq-A) with a separate MULTI-seq library prep workflow performed following cDNA amplification, as described previously (McGinnis, Patterson, et al., Nature Methods, 2019)