Abstract
Cinnamyl alcohol dehydrogenase (CAD) (EC 1.1.1.195) catalyses the final step in lignin precursor synthesis reducing the cinnamyl aldehydes ( para-coumaryl, coniferyl and sinapyl aldehydes) to the corresponding alcohols in the presence of NADPH. In this paper, we report the molecular cloning and characterisation of a Eucalyptus globulus genomic fragment encoding CAD2, and the corresponding full-length cDNA isolated from young stem material. This was achieved using the polymerase chain reaction-based method known as rapid amplification of cDNA ends, with oligonucleotide primers corresponding to regions of homology between CAD-encoding sequences from other eucalypt species. The identity of the clones was inferred by sequence data comparison and the cDNA sequence (1423 bp) was found to encode a protein of 356 amino acid residues. The CAD2 transcript was most abundant in stem, followed by root and midrib tissues, which corresponds with the role of lignin in water retention in plants and in providing mechanical support. Low level expression was also observed in leaf tissue. Southern blot analysis revealed a single CAD gene in this species with the presence of possibly different allelic representations.
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