Petunia hybrida ovule-specific cDNA libraries were screened for genes encoding homologues of protein kinases known to be involved in developmental processes. Two cDNAs, PSK4 and PSK6 (Petunia hybrida shaggy related protein kinase), homologous to GSK-3 from rat, MDS1 from yeast and to the segment polarity gene shaggy/zeste-white 3 (sgg/zw3) from Drosophila were characterized in detail. Southern blot analysis showed that the PSK4 and PSK6 genes belong to a small multigene family in P. hybrida. The PSK4 and PSK6 proteins, which are 70% identical to sgg/zw3 and its functional homologue GSK-3 over the protein kinase catalytic domain and 55% identical to MDS1, represent two different subgroups of protein kinases. RNA gel blot and RT-PCR analyses revealed that the PSK4 gene is expressed during the vegetative phase and the female reproductive phase of development and the PSK6 gene predominantly during the male and female reproductive phases. In situ hybridization on developing flower buds confirmed that PSK4 is expressed throughout the different developmental stages from ovule primordium formation until embryo sac maturation, while PSK6 showed expression during anther cell differentiation and at pollen maturity, and an expression pattern similar to PSK4 in the female reproductive organs. The results also revealed transcripts of a PSK gene in embryos at the globular stage.