An increasing number of studies report that functional divergence in duplicated genes is accompanied by gene expression changes, although the evolutionary mechanism behind this process remains unclear. Our genomic analysis on the yeast Saccharomyces cerevisiae shows that the number of shared regulatory motifs in the duplicates decreases with evolutionary time, whereas the total number of regulatory motifs remains unchanged. Moreover, genes with numerous paralogs in the yeast genome do not have especially low number of regulatory motifs. These findings indicate that degenerative complementation is not the sole mechanism behind expression divergence in yeast. Moreover, we found some evidence for the action of positive selection on cis-regulatory motifs after gene duplication. These results suggest that the evolution of functional novelty has a substantial role in yeast duplicate gene evolution.