Isolation of genes involved in secondary metabolism from Melaleuca alternifolia (Cheel) using Expressed Sequence Tags (ESTs)

Melaleuca alternifolia (Cheel) is a commercially important, cultivated Australian native tree, harvested for its monoterpene-rich essential oil. To isolate genes involved in terpene biosynthesis from this tree, 454 randomly selected clones from a cDNA library derived from flush growth were sequenced and analysed. The five most abundant sequences included cDNAs that encoded Rubisco and photosystem II, as expected. The remaining three abundant sequences included a GATA-binding transcription factor homolog, a sequence with low similarity to a hypothetical protein from a plastid genome of evening primrose (Oenothera elata) and a short sequence with no similarity. These latter three pose interesting targets for functional genomic studies, as their function is unknown and their relative high abundance infer a pivotal role in young tissue of M. alternifolia. Only three ESTs involved in terpene biosynthesis were isolated, a cycloartenol synthase-like protein, a limonene synthase-like protein and farnesyl pyrophosphate synthase, of which only the latter two are involved in the production of the valuable essential oils. However, five ESTs were isolated for phenylpropanoid biosynthesis, excluding those that are committed to lignin biosynthesis. Other secondary metabolite-related ESTs included genes for carotenoid and plant growth regulator biosynthesis, as well as various cytochrome p450s of unknown function. A total of 5% of ESTs isolated in this study are involved directly with secondary metabolite production. This study illustrates the usefulness of ESTs as a method for bio-prospecting for secondary metabolites in simple plant tissues.