Fig. 5. The transcriptional program during mouse preimplantation development. (A) Actinomycin D completely blocked development. Two-cell-stage embryos were treated with actinomycin D (0.04 µM or 0.4 µM), and compared with control embryos (0 µM) at 24 (E2.5) and 48 (E3.5) hours after treatment. (B) Schedule of inhibitor treatment and of the microarray experiment. Solid and broken lines indicate the duration of U0126 and DMSO (vehicle) treatment, respectively. The inhibitor was added at the late two-cell stage, and embryos were collected for microarray experiments at the time points shown with red circles. (C) The numbers of genes whose expression levels in cont. 2.5 or cont. 3.5 were increased or decreased with statistical significance by more than threefold as compared with those in cont. 1.5 are shown. Because overlapping genes exist, in total, 6863 probe sets were differentially expressed. (D) Hierarchical clustering analysis showed the similarity in transcription profiles among the samples tested. This was performed by GeneSpring 7.3. dpc, days post-coitum; cont. 1.5/cont. 2.5/cont. 3.5, control embryos collected at day 1.5/2.5/3.5, respectively; U2.5, U0126-treated embryos collected at day 2.5; U3.5/U4.5, embryos released from the U0126-induced arrest, collected at day 3.5/4.5, respectively.