All genes in this study were mapped on the Arabidopsis genome, and linked to large regions of within genome colinearity. These regions are remnants of a complete genome duplication in the evolutionary history of Arabidopsis, estimated to have occurred 24-86 million years ago (Simillion et al., 2002; Blanc et al., 2003; Bowers et al., 2003; Ermolaeva et al., 2003; Raes et al., 2003).

A complex history of gene duplications caused the expansion and diversification of the respective gene families. Interestingly, the polyploidy event, did not create new classes within any of the investigated families. Mapping of the genome duplication on the respective phylogenetic trees shows that, in all cases, this event together with several small scale duplications, was responsible only for a greater within class diversity. Classes must have originated at an earlier time in evolution, i.e., before 24-86 million years ago and not necessarily all at the same time.

By definition, the complete genome duplication created a full, redundant, double set of monolignol biosynthesis genes. In some gene families (PAL, CAD, CCR, 4CL, and CCoAOMT), duplicates that originated through this event have been retained, whereas they were lost in others (COMT, HCT, and C4H). The mechanisms and reasons of gene conservation and loss after duplication are still unclear and various theories exist (Prince and Prickett, 2002). Some of the duplicated genes in this study that were retained have evolved different expression patterns after the genome duplication (CCR1 and CCR2, PAL3 and PAL4, and CAD1 and CAD7/CAD8), whereas others show no clear difference in expression (PAL 1 and PAL 2). In some cases, the influence of the genome duplication is blurred by more ancient (4CL1, 4CL2, and 4CL4) or recent (CCoAOMT2, CCoAOMT5, and CCoAOMT6) tandem duplications. The observed differences in expression of the CCR and CAD families might point to a functional divergence of these genes, a process called subfunctionalization. The duplicates still exert the same biochemical function, but in different spatio temporal "niches" in the organism (Piatigorsky and Wistow, 1991; Hughes, 1994; Force et al., 1999). A putative example of subfunctionalization after gene duplication is found in the C3H family, where C3H2 and C3H3 show a mutually exclusive expression pattern during stem development. However, although subfunctionalization may be a possible reason why the duplicated genes are retained, only further functional studies will reveal the full consequences of these gene and genome duplications within the monolignol biosynthetic pathway and provide some more hints on the evolutionary constraints acting on these families.