Single cell RNA Sequencing Analysis of Gene Expression Profile in Dgcr8 Conditional KO Embryonic Hearts

Purpose: To compare the single cell transcriptome of wild type and Mesp1Cre+/Dgcr8-/- embryonic heart at embryonic stage E9.5 Methods: For single cell sample preparation, Ventricular part of E9.5 heart tubes were dissected and digested into single cells by 0.04% Trypsin combined with 0.05% Collagenase IV, and then transferred into DMEM containing 10% fetal bovine serum for termination. After washed in PBS without Ca2+, the single cells were manually transferred into cell lysis buffer with a mouth pipette. Total RNA was reverse transcribed and amplified using Smart-seq2 protocol (Picelli et al., 2014), followed by Tn5 tagmentation and PCR enrichment to generate libraries using TruePrep DNA Library Prep Kit V2 for illumina (Vazyme, TD501-503) according to the manufacturer’s suggested protocol. Clean reads were mapped to mouse genome (mm10) using Salmon. Results: Genes significantly differentially expressed in the E9.5 Dgcr8 cKO single cardiomyocyte compared with wild type group were identified. Conclusions: Dgcr8 cKO caused important gene expression changes in E9.5 cardiomyocytes at single cell level Perform single cell RNA-seq of wild type and Mesp1Cre+/Dgcr8-/- embryonic heart at embryonic stage E9.5