# Molecular cloning, prokaryotic expression and induction characteristics of the sesquiterpene synthase gene (AsSS15) from the Chi-Nan germplasm (Aquilaria sinensis)

## TL;DR

This study identifies and characterizes a gene involved in the production of valuable sesquiterpenoids in a high-yielding type of Aquilaria sinensis, which is used to make agarwood.

## Contribution

The study reports the molecular cloning and functional analysis of the AsSS15 gene in Chi-Nan germplasm for the first time.

## Key findings

- The AsSS15 gene is highly expressed in stem and root tissues, key sites for agarwood formation.
- AsSS15 catalyzes FPP to produce nerolidol, confirming its role in sesquiterpene biosynthesis.
- Chi-Nan germplasm shows higher AsSS15 expression compared to ordinary A. sinensis germplasms.

## Abstract

Sesquiterpenoids are one of the secondary metabolites in agarwood which, is a profitable traditional Chinese medicine and spice. Agarwood is derived from the injured Aquilaria sinensis. Chi-Nan germplasm is a new chemotype of A. sinensis that exhibits a higher capacity of agarwood production and the diversity of sesquiterpenoid biosynthesis. To examine the mechanism of sesquiterpene generation from Chi-Nan germplasm, a 1812 bp-length sesquiterpene synthase 15 (AsSS15) gene encoding 603 amino acids was obtained from the wounded branches. AsSS15 showed significant expression levels in the stem and root that the major tissues of agarwood production, indicating that it was involved in agarwood formation. AsSS15 expression was higher in wounded branches of Chi-Nan germplasms than that in ordinary germplasms of A. sinensis. The AsSS15 recombinant protein was successfully expressed, and catalyzed farnesyl pyrophosphate (FPP) substrates to generate nerolidol in vitro. These findings demonstrated that AsSS15 significantly induces the expression of gene related to sesquiterpene biosynthesis in Chi-Nan germplasm. This study first elucidates potential mechanisms of sesquiterpene generation from Chi-Nan germplasms, which adds to the theory of sesquiterpene accumulation in A. sinensis and serves as a foundation for future research into the biosynthesis and application of Chi-Nan germplasm.

The online version contains supplementary material available at 10.1007/s12298-025-01640-z.