Extended Data Fig. 1: Viral vector-mediated transduction and live DNA staining approaches in early mouse embryos and HEK 293 T cells.

a,b Representative images of mouse embryos transduced with a pEGIP lentiviral vector that carries an EGFP gene downstream of the EF-1a promoter. Embryos were washed tree times with hyaluronidase for 10 s and cultured in global media. The zona pellucida of mouse embryos was removed prior to transduction. Mouse zona-free embryos were then divided into two groups for transduction at the: (a) two cell stage or at the (b) morula stage. Embryos were culture in Global medium containing pEGIP lentiviral vectors or in control Global medium in the absence of lentivirus. Embryos were then imaged live 24 h following transduction. c, Transduction of HEK 293 T cells with concentrated pEGIP lentivirus was performed as a control confirming efficacy of EGFP lentiviral vectors. d,e CellLight™ Histone H2B-GFP, BacMam 2.0 baculovirus transduction of early mouse embryos from the (d) 2-cell stage or the (e) morula stage. H2B-GFP BacMam transduced mouse embryos were imaged 24 hrs after transduction. f, Transduction of HEK 293 T cells with H2B-GFP BacMam was performed as a control confirming GFP expression. g, Schematic of AAV6-mediated transduction of embryos. h,i Mouse 2-cell stage embryos were cultured in the presence of AAV6 adeno-associated virus carrying a tdTomato gene downstream of the EF-1a promoter. The expression of tdTomato was analyzed by live cell imaging at the (h) morula or (i) blastocyst stage. Data from three independent experiments. j, Schematic representation of 8-cell stage mouse embryos cultured in Global medium containing live dyes for 24 or 48 hrs until the blastocyst stage. Mouse embryos were subsequently imaged on a confocal microscope in the presence of the dyes. k, Representative images of embryos stained with SPY650-DNA after 24 hrs and 48 hrs and phase-contrast images. Embryos were stained from the 8-cell stage until the times indicated. l, Representative images of embryos stained with various live DNA dyes (SPY650-DNA, 5-TMR-Hoechst, 4-TMR-Hoechst, 4-580CP-Hoechst, 5-580CP-Hoechst, and NucLight Rapid Red). Imaging was conducted 48 hrs post-staining. Phase contrast images are shown. Data from three independent experiments (k-l). Scale bar, 30 μm.