This time-lapse series illustrates the early division up to the four-cell stage of a C. elegans embryo depleted of cdc-48 and atl-1 through RNAi. The first C. elegans embryonic division of the P0 zygote is asymmetric and generates an anterior AB cell, and a smaller posterior P1 cell. These cells have different developmental fates and division timing, with AB dividing approximately 2 min before P1. Time-lapse differential interference contrast (DIC) microscopy identified that downregulation of cdc-48, ufd-1, and npl-4 increases the cell division delay of P1 in comparison to AB, leading to a prolonged three-cell stage. Downregulation of atl-1 suppressed the P1 cell division delay phenotype of cdc-48, ufd-1, and npl-4 RNAi embryos, suggesting that the DNA replication checkpoint is responsible for the delayed P1 cell division. The series corresponds to experiments shown in Fig. 2A and Movie 6 of Mouysset et al. Movie is CIL 25633 and original data file is CIL 35147.
RNA interference was performed using the feeding method. L4 larvae were placed on IPTG-containing plates seeded with Escherichia coli [HT115(DE3)] expressing double-stranded RNA. Eggs were extruded in M9 buffer from dissected adult worms and mounted on 2% agarose pads. Recordings were acquired at 4-s intervals with AxioCam HR or AxioCam MRc cameras mounted on Axioplan2 Imaging or Axiophot microscopes, respectively, equipped with differential interference contrast (DIC) optics (100×/1.3 Plan-Neofluar; Carl Zeiss).
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