Figure 1

Identification of histone arginine residues for early embryonic development. (a) Schematic diagram of the experimental procedures. (b) Development of H2AX-EGFP—(blue bar with round box), H2AXR3A-EGFP—(blue bar with square box), H3.3-EGFP—(green bar with round box), H3.3R2A-EGFP—(green bar with triangular box), H3.3R8A-EGFP—(green bar with square box), H4-EGFP—(red bar with round box), and H4R3A-EGFP—(red bar with square box) expressed embryos. A chi-squared test was performed for analysing embryonic development through to the blastocyst stage. The asterisk indicates statistical significance (***p < 0.001). (c) Representative images of H3.3-EGFP-expressed, H3.3R2A-EGFP-expressed, and untreated embryos at 24 hpi and 96 hpi. Scale bar = 100 µm. (d) Immunofluorescent images of H3R2me2s (red) and DAPI (blue) in H3.3-EGFP-expressed, H3.3R2A-EGFP-expressed, and untreated zygotes. EGFP (green) shows successful injection. Key: ♂, male pronuclei; ♀, female pronuclei. Scale bar = 20 µm. (e) Quantification of H3R2me2s signal intensities in the pronuclei of H3.3-EGFP-expressed, H3.3R2A-EGFP-expressed, and untreated zygotes. Each dot plot represents a single zygote. Red bars indicate the mean values. The mean value of male pronuclei in untreated zygotes was set as 1. One-way ANOVA and the Tukey–Kramer test were performed to evaluate statistical significance. *Significantly different from the control (**p < 0.01, ***p < 0.001); n.s.: Not significantly different from the control. The number of zygotes was 15 for the H3.3-EGFP group, 15 for the H3.3R2A-EGFP group, and 13 for the untreated group. The pronucleus used for quantification are presented in Fig. S5.