GC-MS Analysis
GC-MS analysis was performed via splitless injection using an Agilent 7890A GC system and Agilent Technologies 5975C Inert XL Mass Selective Detector equipped with an HP-5MS UI column (30 m × 0.25 mm × 0.25 μm; Agilent Technologies). Injector temperature was 250°C. Temperature was initially held for 3 min at 40°C, then increased to 130°C at the rate of 10°C·min-1, increased to 250°C at the rate of 50°C/min, and maintained for 10 min; helium gas at a flow of 1 mL/min was used as vector.
MS conditions were as follows: ionization energy, 70 eV; electronic impact ion source temperature, 200°C; quadrupole temperature, 150°C; and mass range, 35-550u. Products were identified based on their retention time and electron ionization mass spectra using mass spectra from the National Institute of Standards and Technology (NIST) Mass Spectral Library (NIST-14.0).
Insect behaviour
For aphids (Myzus persicae ), a leaf disk bioassay was performed as described by Picard et al . (2012). Transformed N. benthamiana leaves were punched into disks of 1 cm in diameter. These disks were placed onto filter paper that was suitable for a Petri dish. Aphids, which had been starved for 3 h, were placed in these Petri dishes, and the number of insects on either control disks or treatment disks was recorded for 1 hour. Ninety aphids were tested in total. We then used a Y-Tube olfactometer to assess the influence of volatiles on ladybugs (Harmonia axyridis ) that had been starved for 3 h. Sixty ladybugs were tested in total. Ladybugs entering into more than half-length of the arms of the olphactometer and lingering for at least 10 s were recorded as responders. Disks were exchanged for fresh disks after 10 aphids or ladybugs were tested.