Abstract
The diseased wood, agarwood, from the tropical tree taxa Aquilaria, is famed for its unique fragrance and medicinal values, mainly due to its richness in secondary metabolites such as the sesquiterpenes. The presence of sesquiterpenes in high numbers and amounts correlates with agarwood of high quality. To understand the synthesis of this important compound, we cloned two candidate genes in the terpenoid synthesis pathway from Aquilaria malaccensis Lam., a major agarwood tree species. The genes encoding sesquiterpene synthase (AmSesTPS1) and δ-guaiene synthase (AmGuaiS1) were successfully cloned from callus RNA, using specific primers derived from transcriptomic data, in a reverse transcription PCR reaction. The full-length complementary DNA (cDNA) sequence of AmSesTPS1 was 1632 bp encoding for 544 amino acids, and AmGuaiS1 was 1644 bp encoding for 547 amino acids. Sequence alignment analysis showed that AmSesTPS1 shared between 99 to 100 % identity with sesquiterpene synthase from Aquilaria sinensis (Lour.) Spreng. while AmGuaiS1 shared between 95 to 99 % identity with δ-guaiene synthases from Aquilaria crassna Pierre ex Lecomte and A. sinensis. The genes were functionally characterised in a time course wounding experiment using 3-year-old living trees. Two types of wood samples were collected: (1) from wounded area (S1) and (2) from 5 cm below the wounded area (S2). AmSesTPS1 was highly expressed after 6 h post wounding for both S1 and S2, at a level three- to six-fold higher than that of the unwounded control (0 h), while AmGuaiS1 was induced after just 2 h of wounding (18- and 5.5-fold at S1 and S2, respectively), after which the expression of both genes was lowered. The average normalised expression of both genes at S1 and S2 indicates the genes were also upregulated in the distal area from the wounding site. It can be deduced that wounding triggers these two genes in the sesquiterpene synthesis pathway, which ultimately leads to agarwood formation.
Highlights
The diseased wood, agarwood, from the tropical tree taxa Aquilaria, is famed for its unique fragrance and medicinal values, mainly due to its richness in secondary metabolites such as the sesquiterpenes
terpene synthases (TPSs)-encoding genes of partial lengths were amplified using primers derived from sequences of A. malaccensis transcriptome (Siah et al 2016) that matched with high homology to the corresponding candidate genes
Polymerase chain reaction (PCR) products of 932 and 1352 bp for AmSesTPS1 and AmGuaiS1, respectively, were obtained and their identities determined by BLASTX searches against the GenBank database
Summary
The diseased wood, agarwood, from the tropical tree taxa Aquilaria, is famed for its unique fragrance and medicinal values, mainly due to its richness in secondary metabolites such as the sesquiterpenes. Aquilaria malaccensis Lam. is the first agarwoodproducing tree listed in the Appendix II of Convention on International Trade in Endangered Species of Wild Fauna and Flora in 1994 (CITES 1994). This tropical tree is of high economic interest because the agarwood it produces is being traded internationally. Agarwood extracts are rich with numerous types of sesquiterpenes such as guaiene, eudesmane, agarospiral and jinkoh-eremol (Yagura et al 2003; Naef 2011). Several important terpenoids in modern medicines are synthesised via similar pathway such as the antimalarial agent artemisin in Artemisia annua L. and the Indian ginseng withanolides in Withania somnifera (L.) Dunal (Rasool and Mohamed 2015)
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