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Fig. 3 Her6 expression dynamics in single cells are altered when the protein is destabilised. (A) Schematic of the Her6:Venus-PEST (HVP) knock-in design. CRISPR/Cas9-mediated HDR resulted in the insertion of the KI cassette including Venus and PEST into the endogenous locus. Detailed description is presented in Fig. S3A,B. (B) Representative examples of Venus oscillatory (HVP-oscillatory) and Venus non-oscillatory (HVP-aperiodic) and their corresponding H2B-mKeima time-series obtained from HVP embryos at 20-30 hpf. (C) Comparison of time-series CV from raw single-cell Venus and H2B-mKeima observed in HV versus HVP. ns: P=0.9381. (D) Relative trend of Venus and H2B-mKeima raw time-series in HV versus HVP embryos. Values >1 indicate upregulation, values=1 indicate steady expression and values <1 indicate downregulation. Circles represent individual cells. ns: P=0.4207. (E) Peak-to-trough fold-change in oscillators expressing raw Venus and their respective mKeima compared in HV and HVP embryos. ns: P=0.4798. (F) Percentage of oscillators in HV versus HVP embryos. Cells expressing Venus (HV and HVP, respectively) and mKeima (H2B-mKeima from the respective HV and HVP embryos). ns: P=0.3609. (G) Coherence values observed in Venus and mKeima compared between HV and HVP time-series. Dots represent individual cells, line indicates median. ns: P=0.8239. (H,I) Period and peak-to-trough fold-change of oscillatory HV versus HVP expression. ns: P=0.3165. (C-I) Boxes indicate median with interquartile range. Dots represent median per embryo in C,E-I. (J) Diagram representation of the main features of HVP oscillations. *P≤0.05; **P≤0.01; ***P≤0.001; ****P≤0.0001 (statistical test details found in Table S6). ns, not significant.