GSM8328864
Forty zebrafish tail samples collected at 24, 38 and 48 hpf, were dissected between the yolk ball and extension and dissociated according to a modified version of the protocol described by Bresciani et al. (2018). Briefly, the tails were dissociated with a mixture of DNaseI (20µg/mL), Collagenase/Dispase (8 mg/mL), and 0.25% Trypsin-EDTA at 30.5°C for 15 minutes. The proteases were quenched with DMEM + 10% FBS. Samples were washed and resuspend in PBS. Cells from each timepoint were hashed with a unique lipid modified oligo using Multi-seq. The barcoded samples were subsequently pooled, washed with 1% BSA + PBS, and resuspended in 0.1% BSA + 18% Optiprep in PBS at a final concentration of ~300,000 cells/mL. Single-cell transcriptomes were captured using inDrops. NGS libraries were prepared by the Single-cell Core at Harvard Medical School and sequenced using an Illumina NovaSeq kit.