title: Single-cell RNA-seq of Mus musculus: adult male colon
design: Single-cell suspensions were obtained from colon tissue of C57BL/6 mice fed with inulin-enriched (10%) diet or a control (AIN93M) diet for 3-4 weeks. The colon was opened longitudinally and washed in cold PBS to remove the feces. Tissue was cut  into small pieces, and  incubated with HBSS-10 mM EDTA-1% P/S for 20 min in a termomixer with 1000 rpm at 37 C. Colon fragments were removed and centrifuged (300 g, 5 min, 4C). The pellet was then washed with cold HBSS-0.04% -1% P/S  and centrifuged again. The supernatant was removed and cells were resuspended in HBSS-0.04% BSA-1% P/S and 10 mM DTT and incubated in a shaker at RT for 10 min. Cold PBS-0.04% BSA was added next, and the suspension was filtered using a 70 um cell strainer. After centrifuging (300 g, 5 min, 4 C), the supernatant was discarded and warm TrypLE Express (37 C) + DNAse 50ug/mL were added to the cells (37 C for 5 min). HBSS with 0.04% BSA and 1% P/S was added to stop the reaction. Cold PBS-0.04% BSA was added and the suspension was filtered (40 um cell strainer). The tubes were spined (300 g, 5 min, 4 C) and the supernatant was removed. Cells were resuspended in cold PBS-0.04% BSA. Single-cell suspensions were counted and then loaded onto the Chromium Next GEM Chip G (10X Genomics) according to recommendations in the manufacturer's protocol CG000388. The quality control of libraries was also performed by the Bioanalyzer high-sensitivity chip. The libraries were sequenced on an Illumina HiSeq with read length of PE150 bp.