AbstractMethane (CH4) formation by vegetation has been studied intensively over the last 15 years. However, reported CH4emissions vary by several orders of magnitude, thus making global estimates difficult. Moreover, the mechanism(s) for CH4formation by plants is (are) largely unknown.Here, we introduce a new approach for making CH4formation by plants clearly visible. By application of13C‐labeled dimethyl sulfoxide (DMSO) onto the leaves of tobacco plants (Nicotiana tabacum) and Chinese silver grass (Miscanthus sinensis) the effect of light and dark conditions on CH4formation of this pathway was examined by monitoring stable carbon isotope ratios of headspace CH4(δ13C‐CH4values).Both plant species showed increasing headspace δ13C‐CH4values while exposed to light. Higher light intensities increased CH4formation rates inN. tabacumbut decreased rates forM. sinensis. In the dark no formation of CH4could be detected forN. tabacum, whileM. sinensisstill produced ~50% of CH4compared to that during light exposure.Our findings suggest that CH4formation is clearly dependent on light conditions and plant species and thus indicate that DMSO is a potential precursor of vegetative CH4. The novel isotope approach has great potential to investigate, at high temporal resolution, physiological, and environmental factors that control pathway‐specific CH4emissions from plants.