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. 2021 Feb 5;12:841. doi: 10.1038/s41467-021-21020-x

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 6 — a Three-dimensional time-lapse images of pronuclei and the cell surface relative to pronuclear formation (0 h) in zygotes injected with 3 μM MBP (control) or FH2 until nuclear envelope breakdown (NEBD). Female (♀) and male (♂) genomes are shown. Phases with respective movement speeds are classified as in Fig. 1a. The mean distance (thick line) of male (b) and female (c) pronuclei centroids to zygote centre during pronuclear migration in zygotes injected with MBP or FH2 was calculated from (a). The total number of analyzed zygotes specified in italics was pooled from three independent experiments. S.d. shown as shaded areas. Statistical plots of average velocities of male (d) and female (e) pronuclei during pronuclear migration in zygotes injected with MBP or FH2 calculated from (b) and (c) with the same sample sizes as in (b) and (c), respectively. Statistical plots show mean ± s.d. Two-tailed Student’s t test was used to test for significance (from left to right: d p < 0.0001, p = 0.5, p = 0.4, p < 0.0001, p < 0.0001 and p = 0.0003; e p = 0.01, p < 0.0001, p = 0.01, p = 0.5, p < 0.0001 and p < 0.0001). f Distance of pronuclei centroids to zygote centre at NEBD for zygotes injected with MBP or FH2 calculated from data sets in (a). Box plot showing median (line), mean (small square), 5th, 95th (whiskers) and 25th and 75th percentile (box). Two-tailed Student’s t test was used to test for significance (male p = 0.4 and female p = 0.5). g Three-dimensional time-lapse images of with cortical mClover-Spire2 and female pronuclei in DMSO- or nocodazole-treated zygotes relative to pronuclear formation (0 min) in live zygotes (z-projection of 20 sections, every 1.5 µm). Second polar bodies (PB) are labelled. h Super-resolution (airyscan) time-lapse images of mScarlet-Spire2 (white) and F-actin (EGFP-UtrCH, pseudocolour, 12-Bit grayscale) in live zygotes treated with DMSO or nocodazole in proximity of the forming female pronucleus displayed as single plane. Time point 0 marks the start of acquisition. Corresponding look-up table is shown as a colour scale bar ranging from intensity levels 1 (low) to 4096 (high). The brighter beans radiating from the cytokinetic furrow (white arrow) of the second polar body extrusion in DMSO-treated zygotes are actin filaments located in the spindle remnant (yellow arrow) and appear distinct from the places that emerge below the cell surface in nocodazole-treated zygotes. Scale bars, 10 µm.