Many aromatic plant volatile compounds contain methyleugenol, which is an attractant for insect pollination and has antibacterial, antioxidant, and other properties. The essential oil of Melaleuca bracteata leaves contains 90.46% methyleugenol, which is an ideal material for studying the biosynthetic pathway of methyleugenol. Eugenol synthase (EGS) is one of the key enzymes involved in the synthesis of methyleugenol. We recently reported two eugenol synthase genes (MbEGS1 and MbEGS2) present in M. bracteata, where MbEGS1 and MbEGS2 were mainly expressed in flowers, followed by leaves, and had the lowest expression levels in stems. In this study, the functions of MbEGS1 and MbEGS2 in the biosynthesis of methyleugenol were investigated using transient gene expression technology and virus-induced gene silencing (VIGS) technology in M. bracteata. Here, in the MbEGSs genes overexpression group, the transcription levels of the MbEGS1 gene and MbEGS2 gene were increased 13.46 times and 12.47 times, respectively, while the methyleugenol levels increased 18.68% and 16.48%. We further verified the function of the MbEGSs genes by using VIGS, as the transcript levels of the MbEGS1 and MbEGS2 genes were downregulated by 79.48% and 90.35%, respectively, and the methyleugenol content in M. bracteata decreased by 28.04% and 19.45%, respectively. The results indicated that the MbEGS1 and MbEGS2 genes were involved in the biosynthesis of methyleugenol, and the transcript levels of the MbEGS1 and MbEGS2 genes correlated with the methyleugenol content in M. bracteata.
Keywords: Melaleuca bracteata; VIGS; biosynthesis; eugenol synthase; methyleugenol; transient overexpression.