Project description:Indian Sandalwood, Santalum album L. is highly valued for its fragrant heartwood oil and is dominated by a blend of sesquiterpenes. Sesquiterpenes are formed through cyclization of farnesyl diphosphate (FPP), catalyzed by metal dependent terpene cyclases. This report describes the cloning and functional characterization of five genes, which encode two sesquisabinene synthases (SaSQS1, SaSQS2), bisabolene synthase (SaBS), santalene synthase (SaSS) and farnesyl diphosphate synthase (SaFDS) using the transcriptome sequencing of S. album. Using Illumina next generation sequencing, 33.32 million high quality raw reads were generated, which were assembled into 84,094 unigenes with an average length of 494.17 bp. Based on the transcriptome sequencing, five sesquiterpene synthases SaFDS, SaSQS1, SaSQS2, SaBS and SaSS involved in the biosynthesis of FPP, sesquisabinene, β-bisabolene and santalenes, respectively, were cloned and functionally characterized. Novel sesquiterpene synthases (SaSQS1 and SaSQS2) were characterized as isoforms of sesquisabinene synthase with varying kinetic parameters and expression levels. Furthermore, the feasibility of microbial production of sesquisabinene from both the unigenes, SaSQS1 and SaSQS2 in non-optimized bacterial cell for the preparative scale production of sesquisabinene has been demonstrated. These results may pave the way for in vivo production of sandalwood sesquiterpenes in genetically tractable heterologous systems.
Project description:Santalum album L. (Indian sandalwood) is an economically important plant species because of its ability to produce highly valued perfume oils. Little is known about the mechanisms by which S. album adapts to low temperatures. In this study, we obtained 100,445,724 raw reads by paired-end sequencing from S. album leaves. Physiological and transcriptomic changes in sandalwood seedlings exposed to 4 °C for 0-48 h were characterized. Cold stress induced the accumulation of malondialdehyde, proline and soluble carbohydrates, and increased the levels of antioxidants. A total of 4,424 differentially expressed genes were responsive to cold, including 3,075 cold-induced and 1,349 cold-repressed genes. When cold stress was prolonged, there was an increase in the expression of cold-responsive genes coding for transporters, responses to stimuli and stress, regulation of defense response, as well as genes related to signal transduction of all phytohormones. Candidate genes in the terpenoid biosynthetic pathway were identified, eight of which were significantly involved in the cold stress response. Gene expression analyses using qRT-PCR showed a peak in the accumulation of SaCBF2 to 4, 50-fold more than control leaves and roots following 12 h and 24 h of cold stress, respectively. The CBF-dependent pathway may play a crucial role in increasing cold tolerance.
Project description:Sandalwood oil is one of the world's most highly prized essential oils, appearing in many high-end perfumes and fragrances. Extracted from the mature heartwood of several Santalum species, sandalwood oil is comprised mainly of sesquiterpene olefins and alcohols. Four sesquiterpenols, ?-, ?-, and epi-?-santalol and ?-exo-bergamotol, make up approximately 90% of the oil of Santalum album. These compounds are the hydroxylated analogues of ?-, ?-, and epi-?-santalene and ?-exo-bergamotene. By mining a transcriptome database of S. album for candidate cytochrome P450 genes, we cloned and characterized cDNAs encoding a small family of ten cytochrome P450-dependent monooxygenases annotated as SaCYP76F37v1, SaCYP76F37v2, SaCYP76F38v1, SaCYP76F38v2, SaCYP76F39v1, SaCYP76F39v2, SaCYP76F40, SaCYP76F41, SaCYP76F42, and SaCYP76F43. Nine of these genes were functionally characterized using in vitro assays and yeast in vivo assays to encode santalene/bergamotene oxidases and bergamotene oxidases. These results provide a foundation for production of sandalwood oil for the fragrance industry by means of metabolic engineering, as demonstrated with proof-of-concept formation of santalols and bergamotol in engineered yeast cells, simultaneously addressing conservation challenges by reducing pressure on supply of sandalwood from native forests.
Project description:Santalum album (sandalwood) is one of the economically important plant species in the Santalaceae for its production of highly valued perfume oils. Sandalwood is also a hemiparasitic tree that obtains some of its water and simple nutrients by tapping into other plants through haustoria which are highly specialized organs in parasitic angiosperms. However, an understanding of the molecular mechanisms involved in haustorium development is limited. In this study, RNA sequencing (RNA-seq) analyses were performed to identify changes in gene expression and metabolic pathways associated with the development of the S. album haustorium. A total of 56,011 non-redundant contigs with a mean contig size of 618 bp were obtained by de novo assembly of the transcriptome of haustoria and non-haustorial seedling roots. A substantial number of the identified differentially expressed genes were involved in cell wall metabolism and protein metabolism, as well as mitochondrial electron transport functions. Phytohormone-mediated regulation might play an important role during haustorial development. Especially, auxin signaling is likely to be essential for haustorial initiation, and genes related to cytokinin and gibberellin biosynthesis and metabolism are involved in haustorial development. Our results suggest that genes encoding nodulin-like proteins may be important for haustorial morphogenesis in S. album. The obtained sequence data will become a rich resource for future research in this interesting species. This information improves our understanding of haustorium development in root hemiparasitic species and will allow further exploration of the detailed molecular mechanisms underlying plant parasitism.
Project description:Santalum album Linn. is an evergreen and hemi-parasitic tree, the heartwood-sandalwood of which was used during a long history in traditional Chinese medicine. Kuhnia rosmarinifolia Vent. is a good host for 1- or 2-year-old growing S. album. The interaction between S. album and K. rosmarinifolia is still little known. Many studies have been carried out on a number of plants for identification and diversity of endophytes. In this study, in total 25 taxa of endophytic fungi were isolated from the roots of S. album and the roots of K. rosmarinifolia. The most frequently isolated genera were Penicillium sp. 1 and Fusarium sp. 1 in the roots of S. album and K. rosmarinifolia, respectively. S. album is a root parasite of K. rosmarinifolia. The interesting result is that they apparently do not share the same endophytic fungi isolates. This study for the first time explored the content of endophytic fungi from S. album and K. rosmarinifolia, which provides important information for further studies.