Feeding Assay to Study the Effect of Phytocytokines on Direct and Indirect Defense in Maize

Phytocytokines mediate defense against pests and pathogens. Many methods have been developed to study the physiological responses triggered by phytocytokines in dicot plants. Here, we describe a detailed peptide feeding protocol to study the effect of phytocytokines on direct and indirect anti-herbivore defense in maize. This method relies on peptide uptake by the excised maize seedling or leaves via the transpiration stream. The headspace volatiles from plant samples are then analyzed by proton transfer reaction time-of-flight mass spectrometry (PTR-TOF-MS), or by gas chromatography–mass spectrometry (GC-MS). The samples can also be further processed to evaluate phytocytokine-induced defense gene expression or phytohormone production.