GSM4969715: PN8_4dpf_uninjured_mpeg1_mcherry_R2; Danio rerio; RNA-Seq
Muscle injury region was dissected out and tissue dissociated into a single cell suspension. Whole larva were utilised for uninjured time point. Cells were sorted using a FACS Aria II (BD biosciences). Live individual macrophages (based on mCherry fluorescence, DAPI exclusion and forward and side scatter properties) were sorted into pre-prepared 384 well plates containing 100-200 nl of CEL-seq primers, dNTPs and synthetic mRNA Spike-Ins contained in 5 μl of Vapor-Lock (Qiagen). Immediately following sorting, plates were spun down and frozen at -80°C until sequencing. Single-cell RNA-sequencing libraries were prepared using the SORT-seq platform. Here the Cel-Seq2 protocol is followed with the aid of robotic liquid handlers. This protocol results in each cell being barcoded, and generating single-cell transcriptomes of all isolated macrophages.