GSM5795726: DSS-treated colon mucosa, experimental day 23; Mus musculus; RNA-Seq
Colons were dissected, manually stripped of myenteric layers, and enzymatically digested in Liberase TM/DNAse I. Dissociated tissue was strained through 70 and 40-micron filters, and the resulting single-cell suspension was treated with a red blood cell lysis buffer. Dead cells were removed using a magnetic separation kit with antibodies targeted against annexin V. The purified suspension was either frozen in 90% fetal bovine serum supplemented with 10% dimethyl sulfoxide (Batch A), or directly submitted (Batch C) for single-cell analysis and library prep via the 10X Genomics Chromium platform. Samples in Batch A were thawed and processed on the same day, representing a single batch run. Samples in Batch C were directly processed after cell isolation, representing a separate batch run. Cells were washed with PBS and pre-loading viability was confirmed to exceed 75%. Cells were run on the Chromium (10X Genomics) platform for single-cell lysis, reverse transcription, barcoding, and library preparation. Paired-end sequencing of 3' transcript ends was performed on a novaSeq machine (Illumina).