Functional genomics of a living fossil tree, Ginkgo, based on next-generation sequencing technology.

Ginkgo biloba is monotypic species native to China and has old, dioecious, medicinally important characteristics. The functional genes related to these characteristics have not been effectively explored due to a limited number of expressed sequence tags (ESTs) from Ginkgo. To discover novel functional genes efficiently and to understand the development of a living fossil tree, Ginkgo, we used massive parallel pyrosequencing on the Roche 454 GS FLX Titanium platform to generate 64 057 ESTs. The ESTs combined with the 21 590 Ginkgo ESTs in genbank were assembled into 22 304 unique putative transcripts, in which 13 922 novel unique putative transcripts were identified by 454 sequencing. After being assigned to putative functions with Gene Ontology terms, a detailed view of the Ginkgo biological systems was displayed, including characterization of unique putative transcripts with homology to known key enzymes and transcription factors involved in ginkgolide/bilobalide and flavonoid biosynthetic pathways, as well as unique putative transcripts related to development, response to disease and defence. The fact that three full-length Ginkgo genes encoding key enzymes were found and cloned, suggests that high-throughput sequencing technology is superior to traditional gene-by-gene approach in discovery of genes. Additionally, a total of 204 simple sequence repeat motifs were detected. Our study not only lays the foundations for transcriptome-led studies in biosynthetic mechanisms, but also contributes significantly to the understanding of functional genomics and development in non-model plants.