This article reviews several research challenges for GMO detection as some pending questions still exist and need more research efforts to be solved. The question of how to apply validation in a modular scheme is handled. It is followed by the problem of how to manage the detection of an always increasing number of new events and the fact that in such a context screening will probably be more and more important. The issue of detection of unauthorized GMOs with special attention to unknown GMOs is considered. Some technical limitations in result expression with respect to botanical impurities or stacked events are also addressed. Finally the establishment of plasmid reference calibrants as alternative to the current plant-derived certified reference materials, their distribution and utilisation are discussed.