Fig. 4

Expression of miR-29 and miR-200 is dysregulated in gill filament culture under hypertonic stress. Gill filaments were challenged in hyperosmotic (Hyper: 600 mOsmol l^− 1) media for 4 h and 8 h and were subsequently processed for small RNA sequencing. a & b Volcano plot showed differentially expressed miRs under 4 h (a) or 8 h (b) hypertonic treatment in gill filaments compared with the control. The p < 0.05 and |Log2 (fold change)| > 1 were set as threshold for significantly differential expression. In 4 h hypertonic treatment, 81 differentially expressed miRs including 42 up-regulated miRs and 39 down-regulated miRs were identified, while in 8 h hypertonic treatment, 55 differentially expressed miRs including 24 up-regulated miRs and 31 down-regulated miRs were identified. c Venn diagram shows the similarity of differentially expressed miRs under 4 h and 8 h hypertonic treatments. Two miRs, miR-29b-3p and miR-200b-3p, with significant expression changes (p < 0.001) in both hypertonic treatment groups were selected for further analysis. d Bar charts exhibit the differential expression levels of miR-29b-3p and miR-200b-3p in control and hypertonic groups (4 or 8 h) based on deep sequencing data. e qRT-PCR was performed to validate the expression pattern of miR-29b-3p and miR-200b-3p. snRNA U6 was used as internal control for normalization of miRNA expression. Results (mean ± s.e.m.) were from five independent experiments. * P < 0.05, ** P < 0.01 vs control