In animal embryos, transcription is repressed for a definite period of time after fertilization. In the embryo of the ascidian, Ciona intestinalis (type A; or Ciona robusta), transcription of regulatory genes is repressed before the 8- or 16-cell stages. This initial transcriptional quiescence is important to enable the establishment of initial differential gene expression patterns along the animal-vegetal axis by maternal factors, because the third cell division separates the animal and vegetal hemispheres into distinct blastomeres. Indeed, maternal transcription factors directly activate zygotic gene expression by the 16-cell stage; Tcf7/β-catenin activates genes in the vegetal hemisphere, and Gata.a activates genes in the animal hemisphere. In the present study, we revealed the dynamics of Gata.a and β-catenin, and expression profiles of their target genes precisely. β-catenin began to translocate into the nuclei at the 16-cell stage, and thus expression of β-catenin targets began at the 16-cell stage. Although Gata.a is abundantly present before the 8-cell stage, transcription of Gata.a targets was repressed at and before the 4-cell stage, and their expression began at the 8-cell stage. Transcription of the β-catenin targets may be repressed by the same mechanism in early embryos, because β-catenin targets were not expressed in 4-cell embryos treated with a GSK inhibitor, in which β-catenin translocated to the nuclei. Thus, these two maternal factors have different dynamics, which establish the pre-pattern for zygotic genetic programs in 16-cell embryos.