Fig. 6: Pig acrocentric chromosomes have less cohesin than metacentric chromosomes.

a Representative Airyscan immunofluorescence images of the measurements from the sister chromatids in the three different chromosome groups. Magenta, kinetochores, (ACA); grey, TALE (anti-GFP); blue, chromosomes, (Hoechst); Dashed circles indicate the sister kinetochores and the white line indicates the distance measured. Scale bar, 1 μm. b Box plot showing measurements of the interkinetochore distance of sister chromatids on metaphase II intact spindles of porcine oocytes (41 oocytes analysed). Box plot shows the median (horizontal black line), mean (small black squares), 25th and 75th percentile (boxes), and outliers (whiskers). Two-tailed unpaired t-test was performed (acro-labelled/meta, p = 1.41075e − 8; acro-labelled/meta-labelled, p = 7.51899e − 9; meta/meta-labelled, p = 0.31898). c Representative Airyscan immunofluorescence images of the SMC3 staining on chromosomes in metaphase II intact spindles of porcine eggs. Single z-plane images from the three different chromosome groups. Magenta, kinetochores, (ACA); grey, TALE (anti-GFP); blue, chromosomes, (Hoechst); green, cohesin, (anti-SMC3); Scale bar, 1 μm. d Box plot showing mean intensity measurements of SMC3 signal on chromosomes from metaphase II intact spindles of porcine eggs (40 oocytes analysed). Box plot shows the median (horizontal black line), mean (small black squares), 25th and 75th percentile (boxes), and outliers (whiskers). Two-tailed unpaired t-test was performed, (acro-labelled/meta, p = 6.11772e − 7; acro-labelled/meta-labelled, p = 2.04894e − 4; meta/meta-labelled, p = 0.37062). e Representative Airyscan immunofluorescence images of the REC8 staining on chromosomes from metaphase I intact spindles of porcine oocytes. Images from the three different chromosome groups. Images are projections of 16 z-planes. Dashed circles indicate the area of the measurements. Magenta, kinetochores, (ACA); grey, TALE (anti-GFP); blue, chromosomes, (Hoechst); green, cohesion, (anti-REC8); Scale bars, 0.5 μm. f Box plot of mean intensity measurements of REC8 intensity around the pericentromeric areas on chromosomes from metaphase I intact spindles of porcine oocytes (49 oocytes analysed). Box plot shows the median (horizontal black line), mean (small black squares), 25th and 75th percentile (boxes), and outliers (whiskers). Two-tailed unpaired t-test was performed (acro-labelled/meta, p = 3.89493e − 24; acro-labelled/meta-labelled, p = 1.41981e − 13; meta/meta-labelled, p = 0.20548). g Percentage of the first chromosome to separate after Trim-Away of REC8 in metaphase II porcine eggs. h Time of separation for each of the six acro-labelled and meta-labelled chromosomes after Trim-Away of REC8 in porcine eggs. As time point zero in each egg is the time of the first chromosome separation (10 oocytes analysed). Box plot shows the median (horizontal black line), mean (small black squares), 25th and 75th percentile (boxes), and outliers (whiskers). Two-tailed unpaired t-test was performed between each separation event for acro-labelled and meta-labelled (1st separation, p = 0.01326; 2nd separation, p = 0.96525; 3rd separation, p = 0.47748;4th separation, p = 0.94742, 5th separation, p = 0.82915, 6th separation, p = 0.2905). i Still images from a time-lapse movie after Trim-Away of REC8 in metaphase II porcine eggs. Time is indicated in minutes and time point 0 is the first frame when imaging started after the injection of the REC8 antibody. Magenta, Metacentric-TALE (Metacentric-TALE-GFP); green, acrocentric label, (Acrocentric-TALE-mScarlet); blue, chromosomes, (H2B-SNAPf). Scale bar, 1 μm. Number of chromosomes analysed is indicated in brackets under each category. P-values in the graphs are indicated as follows, *p < 0.05, ****p < 0.0001, and n.s.: non-significant. The number of cells is indicated in brackets in the figure legend.