title: GSM8825874: DM-1; Mus musculus; RNA-Seq
library_construction_protocol: The isolated  cells were resuspended in digestion buffer (HBSS with Mg2+/Ca2+supplemented with 2 mg/ ml collagenase type l and 40 U/ml DNase l) and incubated at 37 ℃ for 30 min. Cells were washed and re-suspended in PBS and stained w ith a Viability Dye, washed and resuspended in FACS buffer (HBSSwo, 2% v/v FCS, 5mM EDTA). Alive cells were sorted using a FACS Jazz (Becton Dickinson) according to the presence or absence of mCherry signal using a C57Bl6 mouse as negative control. Cells were washed inHBSS-/- and resuspended in HBSS-/- supplemented with 0.04% w/v BSA. Library was performed according to the manufacter's instructions (single cell 3' v2 protocol, 10x Genomics). Briefly, Cells were resuspended in the master mix and loaded together with partitioning oil and gel beads into the chip to generate the gel bead-in-emulsion (GEM). The poly-A RNA from the cell lysate contained in every single GEM was retrotranscripted to cDNA, which contains an llumina R1 primer sequence, Unique Molecular ldentifier (UMl) and the 10x Barcode. The pooled barcoded cDNA was then cleaned up with Silane DynaBeads, amplified by PCR and the apropiated sized fragments were selected with SPRlselect reagent for subsequent library construction. During the library construction llumina R2 primer sequence, paired-end constructs with P5 and P7 sequences and a sample index were added.