GSM5416739: Heart.scRNA.WT1; Mus musculus; RNA-Seq
Tissue samples were digested with 0.25% trypsin and gently dispersed into single-cell suspension with pipetting. Cells were filtered with a 40 μm cell strainer and subjected to fluorescence-activated cell sorting (FACS) to select for live cardiomyocytes which had positive tdTomato fluorescence and negative DAPI staining. Sorted live cardiomyocytes were then captured with Chromium Next GEM Single Cell 3ʹ Reagent Kits v3.1 (10X Genomics) following manufacturer's instructions. Single cells were partitioned into nanoliter-scale Gel Beads-in-emulsion (GEMs) using Chromium Next GEM Chip G in the chromium controller. Immediately following GEM generation, the Gel Bead was dissolved, primers were released, and co-partitioned cell was lysed, and its mRNA were reverse transcribed into barcoded cDNA. After further cleanup and amplification, the cDNA was enzymatically fragmented and amplified via PCR to generate sufficient mass for library construction. After end repair, A-tailing, adaptor ligation and sample index PCR, the library comprised constructs containing the sample index, UMI sequences, barcode sequences, and Illumina standard sequencing primers P5 and P7 at both ends.