Fig. 3

chch is zygotically expressed prior to the mid-blastula transition. To determine if the early nuclear stain observed represents new zygotic transcripts, embryos were microinjected with the transcription inhibitor Actinomycin D (B) or DMSO (A). Embryos were collected at the 64-cell stage and examined for chch expression by RNA in situ hybridization. Following Actinomycin D treatment, the nuclear stain is lost. As a further test to confirm early zygotic transcription of chch, real-time PCR was performed on 1-cell through 16-cell embryos to determine if the relative amount of chch gene expression is increased as development progresses. This analysis revealed an increase in chch expression over these stages (C). Together these results show that chch is zygotically transcribed prior to the mid-blastula transition.