Extended Data Fig. 9: Further characterization of ruminant iCHI embryos.
From: Generation of modified cows and sheep from spermatid-like haploid embryonic stem cells
a. Representative phase contrast and fluorescence images of ruminant haSCs at 0‐, 24-, 48‐, and 72‐h post‐protamine-induction (top panel). The bottom panel shows higher-magnification images. Note that after 48 h protamine-induction, the cell nuclei acquire a spermatid-like structure, detach from the culture dish, and float in the medium. After 72 h of protamine-induction, the protaminized cells showed signs of degeneration. These haSCs harbour the doxycycline (dox)‐inducible PRM1-EGFP vector. Scale bars: in top, 100 μm; in bottom, 20 μm. b. Quantifications of spermatid-like cells in ruminant haSCs at 0‐, 24-, 48‐, and 72‐h post‐protamine-induction. Data are mean ± s.d. (n = 3 independent experiments); P values are from unpaired, two-tailed Student’s t-tests. c. Heatmap and pileup of CUT&Tag signal for the protamine deposition in b-haSCs at 0‐, 24-, 48‐, and 72-h post‐protamine-induction. Each row of the heatmap is a genomic locus and rows are sorted from highest peak intensity to lowest. d. Representative images of dissected bovine uteri isolated from surrogate mothers after day-65 of iCHI embryo transfer. e. Summary of the morphometric measurements of bovine conceptuses at E75 produced by Pro-iCHI or IVF techniques. Data are mean ± s.d. (n = 5 IVF, n = 2 Normal, n = 12 Retarded; n represents total embryos of three independent experiments); P values are from unpaired, two-tailed Student’s t-tests by comparison with IVF group. f. Representative images of dissected ovine uteri isolated from surrogate mothers after day-65 of iCHI embryo transfer. g. Summary of the morphometric measurements of ovine conceptuses at E65 produced by Pro-iCHI or IVF techniques. Data are mean ± s.d. (n = 5 IVF, n = 17 Retarded; n represents total embryos of three independent experiments); P values are from unpaired, two-tailed Student’s t-tests by comparison with IVF group. h. An image of stillborn ovine Pro-iCHI conceptus isolated from surrogate mother after day-150 of embryo transfer.
