In this paper, a lab-on-chip system dedicated to fungi cultivation in Earth’s gravity and simulated microgravity, being a solution that could be used in future nanosatellite missions, is shown. For the first time, a fully glass lab-on-chip structure enabling the proper environment for cultivation of fungi species—Fusarium culmorum—is presented. Apart from the biological validation of the fungi cultures with the use of the lab-on-chip system, tests were carried out under induced microgravity utilising a Rotary Wall Vessel. Correct functioning of the lab-on-chip system was obtained, enabling the growth of fungi spores both in ground and in simulated microgravity conditions. Interestingly, culturing tests have shown that microgravity stimulates the growth of fungi notably, compared to the ground-based experimentation performed simultaneously. The findings of this study can provide substantial new knowledge on microscopic fungi cultivation in lab-on-chip devices, other soil organisms, as well as a potential behavior of these species in microgravity conditions. Culturing system shown in this work can help mycologists to provide better understanding of microscopic fungi nature and their development mechanisms at a single spore level. This opens the way towards regular usage of microfluidic tools in agriculture and horticulture fields and more importantly, in future research on microscopic fungi in space, e.g., as a part of nanosatellite missions.