title: GSM8027657: MI-stiff snRNA-seq rep2; Rattus norvegicus; RNA-Seq
library_construction_protocol: Nuclei were isolated with Nuclei EZ Lysis buffer (NUC-101; Sigma-Aldrich) supplemented with protease inhibitor (5892791001; Roche) and RNase inhibitor (N2615; Promega and AM2696; Life Technologies). Heart samples were cut into 2-mm pieces and homogenized using a Dounce homogenizer (885302-0002; Kimble Chase)  in 2 ml of ice-cold Nuclei EZ Lysis buffer, and they were incubated on ice for 5 minutes with an additional 2 ml of lysis buffer. The homogenate was filtered through a 40-mm cell strainer (43-50040-51; pluriSelect) and then centrifuged at 5003g for 5 minutes at 4°C. The pellet was resuspended and washed with 4 ml of the buffer, and then, it was incubated on ice for 5 minutes. After another centrifugation, the pellet was resuspended in Nuclei Suspension Buffer (1xPBS, 0.07% BSA, and 0.1% RNase inhibitor), filtered through a 20-mm cell strainer (43-50020-50; pluriSelect), and counted. cDNA libraries were constructed using Chrominum Single-Cell 3'GEM, Library & Gel Bead Kit v3, then sequenced on illumina novaseq 6000.