a A protein phylogenetic tree was constructed of obtained sequences. Zebrafish and other chordate Vps50 protein sequence were identified from the Uniprot database. Accession numbers: Human: Q96JG6; Rat: F1LSG8; Mouse: Q8CI71; Zebrafish: F1R0A4; Chicken: Q5ZKV9; Chimpanzee: H2QUX7; Bovine: F1ML08; Western clawed frog: A9UM56; Japanese pufferfish: H2UPA7. b Expression patterns of zebrafish vps50 during embryogenesis. Expression of vps50 gene was detected around the yolk sac at 1 dpf. Once the embryo reached the hatching period (2–3 dpf), vps50 expression was seen in the head and heart (Right Figure). Representative images of n = 3 replicates where with each replicate having at least 15 embryos. Control images of sense strand probed on early embryos (Left Figure)

a Distribution of the four categories in each of the experimental groups. We clustered these injected embryos into four categories according to the severity of their morphology. Grade 1: WT like; Grade 2: mild, curving axis compared with WT embryos; Grade 3: moderate, up to 1/3 shortened axis; Grade 4: severe, the body axis just extended out of the range of the yolk ball. b Injection of vps50-MOs yielded high proportion abnormal phenotypes. Wild-type VPS50 exhibited higher efficiency saving ability than the p. Gly169Val mutation. Dosage-dependent rescue of vps50 knock-down zebrafish embryo with Human VPS50 mRNA. 6 ng vps50-MOs were coinjected with 0, 200, and 400 pg of VPS50 mRNA. The number above each bar was the total number of embryos counted. The P value was calculated by chi-squared analysis