Arbuscular mycorrhizal fungi (AMF) have been shown to improve the growth, health, nutrient uptake, flowering and drought tolerance of many terrestrial plant species. Green roofs are generally deficient in nutrients, organic matter and water, and therefore AMF could be extremely beneficial in improving green roof plant performance. Despite this there is a lack of empirical research into artificially introducing AMF into green roof substrates. In this study, a commercial AMF inoculum was applied to Prunella vulgaris green roof plugs grown in small modules on a flat roof in Sheffield, UK. The modules were filled with commercial green roof substrate (80% small particle sized crushed brick, 20% green waste compost) to a depth of 100 mm. AMF inoculum was applied as four treatments: (i) directly with plug, (ii) mixed evenly into surrounding substrate, (iii) split between plug and substrate, (iv) control treatment with no inoculum added. Significantly greater levels of AMF colonisation of P. vulgaris roots was detected in all AMF treatments compared to the control. Low levels of AMF colonisation of P. vulgaris roots were also observed in the control treatment, confirming that low levels of AMF inoculum were present in this commercial sub-strate. Shoot phosphorous (P) concentration was improved in all AMF treatments, however there was no significant effect of any AMF treatment on P. vulgaris growth rate or biomass production. The highest AMF colonisation of P. vulgaris roots was observed when AMF inoculum was directly added to just the plug. Promisingly, P. vulgaris flowering time at the end of the first growing season was also extended in the plug AMF treatment only. This study has confirmed that commercial AMF inoculum can be used to successfully colonise plants and introduce AMF networks into green roof substrate. Although AMF inoculum was naturally present in the substrate used in this study, levels were extremely low, and unlikely to have any significant effect on plants. This study indicates that care should be taken in the use of AMF inoculum on green roofs, as the growth and health benefits of AMF are not always immediately apparent for green roof plants. In addition much more research is required in order to fully assess the extent of the benefits of AMF on green roof plants and to determine if their use can be financially viable.