alias: GSM3678222
title: GSM3678222: aMHC-Vegfb p7 WT; Mus musculus; RNA-Seq
library_construction_protocol: Mouse hearts were isolated for single-cell preparation as described previously. Hearts were harvested and coronary arteries were perfused briefly with buffer A (113 mM NaCl, 4.7 mM KCl, 0.6 mM KH2PO4, 0.6 mM Na2HPO4, 1.2 mM MgSO4, 12 mM NaHCO3, 10 mM KHCO3, 10 mM HEPES and 30 mM Taurine) to remove blood in a retrograde manner by cannulating the aorta before Collagenase type II (Worthington) was added to dissociate the ventricular cells. The dissociated cells were then allowed to sediment at 37°C for 10 minutes to allow cardiomyocytes to settle down as a pellet leaving behind a non-myocyte rich supernatant. The non-myocyte enriched supernatant was treated with rat anti-mouse CD16/CD32 (eBioscience, clone 2.4G2) at 4°C for 20 minutes before being incubated with fluorophore-conjugated antibodies at 4°C for 30 minutes. The following antibodies were used: anti-CD45-FITC (Biolegend, clone 30-F11), anti-PDGFR-PECy7 (eBioscience, clone APA5) and anti-CD105-PE (eBioscience, clone MJ7/18). Cell suspensions were triturated through a 40 µm Nylon cell strainer (Falcon) and then subjected to flow cytometry-based sorting using a Biorad S3e cell sorter. Subsequent cDNA purification, amplification (12 cycles) and library construction (sample index PCR 14 cycles) was performed as instructed Sample libraries were sequenced on the Illumina NovaSeq 6000 system using S1 flow cell (Illumina)
identifiers.namespace: GEO
identifiers.id: GSM3678222