GSM9335678: hand2_ko; Mus musculus; RNA-Seq
To isolate total cardiac ECs from Hand2f/f;VE-cad-CreERT2; iSure-Cre mice and iSure-Cre mice, we induced Hand2 deletion at E11.5-12.5, treated the mice with tamoxifen, and isolated the hearts at E14.5. HD-wt (iSure-Cre mice) and HD-ko (Hand2f/f;VE-cad-CreERT2; iSure-Cre) hearts were dissected, minced, and digested in a cell-dissociated solution. The cell pellet was resuspended and incubated with CD31-antibody (BD Pharmingen, 551262). Twice the volume of blocking solution was added to stop the reaction, and centrifugation was then performed. The cells were resuspended in the appropriate blocking solution and stored on ice prior to sorting. DAPI (1:1000) was then added to label the dead cells. A FACSAria III flow cytometer (BD Biosciences) was used for sorting. Approximately 9000-12000 DAPI-, APC-CD31+/Tomato+ ECs were sorted per sample for the subsequent experiments. According to the manufacture's introduction, Single-cell RNA-seq libraries were constructed using Single Cell 3' Library and Gel Bead Kit V3.1. The libraries were finally sequenced using an Illumina Novaseq6000 sequencer with a sequencing depth of at least 30,000 reads per cell with pair-end 150 bp (PE150) strategy.