Figure 4.

Down-regulation of aurora kinase C leads to the two-cell arrest, but does not affect the degradation of maternal mRNAs or zygotic activation of selected genes. Early zygotes were injected with dsRNA for GFP or aurora kinase C. Embryos were cultured in vitro until the blastocyst stage. Images were taken using fluorescence light to identify embryos injected with both dsRNA and rhodamine-conjugated dextran. Embryos of each experimental group were scored according to their developmental progress. The number of embryos in each experiment is indicated as injected embryos (two-cell stage; IE2), arrested embryos (A), embryos that developed to blastocyst (EB), and the number of embryos fragmented or dead (D). (A) RNAi GFP-injected control group (IE2 = 18, EB = 15, D = 3) and RNAi aurora kinase C (Aurkc; IE2 = 20, A = 16, D = 4). RT-PCR analysis was performed 24 h after injection, as shown on the right panel. Aurkc, aurora kinase C. This shows that aurora kinase C mRNA is greatly down-regulated, whereas Ccne2 and Oct4 mRNA remains constant in both experiments. Nested PCR on first-strand cDNA generated from either control or RNAi aurora kinase C two-cell embryos. (B) Mosg, Gbx2, Ccne2, Fgfr2, Lepr2, Psme3, Hhex, p270/ARID1A, Dncic2, Fgf9, and control genes Ago2, Aurkc, actin, tubulin, and Oct4. All genes tested showed similar expression levels of the transcripts as the control, except aurora kinase C, which was greatly reduced in the aurora kinase C-depleted embryos.