The importance of cell separation in plant development cannot be overemphasized. The polygalacturonases (PGs) are the one of cell wall hydrolytic enzyme families that has been associated with various cell separation processes in plant development including seed germination, dehiscence, organ abscission, and fruit ripening. Both Arabidopsis and rice PG gene family have expanded in a lineage-specific fashion after the split more than 150 million years ago. Tandem duplications and large-scale duplications are the major contributors to the current PG family size in Arabidopsis. The spatial expression analysis of the 66 Arabidopsis PG family members have led us to conclude that different duplication mechanisms affect the expression divergence differently. This becomes more apparent when temporal examination of expression is conducted in five developmental stages of floral organ abscission in Arabidopsis. Nine distinct patterns of PGs are identified during floral organ abscission in Arabidopsis. Four PGs are specifically upregulated during abscission associated with cell separation process. Careful understanding of relationships among Arabidopsis PGs in a context of evolution together with expression analysis of these PGs will facilitate the functional study of PGs specifically in floral organ abscission in Arabidopsis.