GSM8058226: CI_1_S1; Mus musculus; RNA-Seq
The cardiac tissue was homogenized and Ter119 antibody (BioLegend, 1:200) was used to exclude blood cells. After sorting, nuclei integrity was assessed on 5µl sample diluted 1:2 in Trypan blue and on the Countess III cell counter (Thermofisher). Each nuclei suspension was loaded into one port of a Chromium Next GEM Chip G (10x Genomics) with a target output of 10,000 nuclei. Single nuclei were encapsulated into emulsion droplets using the Chromium Controller (10x Genomics). SnRNA-seq libraries were prepared using the Chromium Next GEM Single Cell 3' Kit v3.1 (10x Genomics) following the manufacturer instructions and each library was amplified using a SureCycler 8800 thermal cycler (Agilent Technologies). The average size of each library was then calculated using a High sensitivity DNA chip on a 2100 Bioanalyzer (Agilent Technologies) and the concentration was determined using the Qubit fluorometer (Thermofisher). Individual libraries were diluted to 10 nM and pooled for sequencing. Library pool was sequenced at 650 pM in paired-end reads (28bp Read1, 10bp Index1, 10 pb Index 2 and 90bp Read2) on a P3 flow cell (100 cycles) of the NextSeq 2000 (Illumina).