The content of patchouli alcohol in transiently overexpressed P. cablin leaves was detected as previously described (Chen et al., 2019 (link)).
A total of 0.4 g leaves of transgenic tobacco were ground frizzed in liquid nitrogen, extracted with 1.5 ml hexane ultrasonic for 30 min, and then heated in a 56°C water bath for 40 min. The sample was centrifuged at 12,000 rpm/min for 3 min, the supernatant was passed through 0.22 μm organic membranes, and the filtrate was transferred into new vials for GC-MS analysis using an Agilent 7890B Gas Chromatograph with 5977A inert Mass Selective Detector (Agilent, United States). The gas chromatograph was equipped with an HP-5MS capillary column (30 m × 0.25 mm × 0.25 μm film thickness). The oven temperature was programmed as follows: from 35°C (5 min hold) to 300°C at a rate of 12°C/min and hold 5 min at 300°C. NIST14/Wiley275 Mass Spectral Library was used for metabolite identification. To analyze transgenic tobacco metabolites, the relative quantification of volatiles was carried out using the external standard method with cyclohexanone as the external standard substance.
A total of 0.4 g leaves of transgenic tobacco were ground frizzed in liquid nitrogen, extracted with 1.5 ml hexane ultrasonic for 30 min, and then heated in a 56°C water bath for 40 min. The sample was centrifuged at 12,000 rpm/min for 3 min, the supernatant was passed through 0.22 μm organic membranes, and the filtrate was transferred into new vials for GC-MS analysis using an Agilent 7890B Gas Chromatograph with 5977A inert Mass Selective Detector (Agilent, United States). The gas chromatograph was equipped with an HP-5MS capillary column (30 m × 0.25 mm × 0.25 μm film thickness). The oven temperature was programmed as follows: from 35°C (5 min hold) to 300°C at a rate of 12°C/min and hold 5 min at 300°C. NIST14/Wiley275 Mass Spectral Library was used for metabolite identification. To analyze transgenic tobacco metabolites, the relative quantification of volatiles was carried out using the external standard method with cyclohexanone as the external standard substance.
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