The single surface glycoprotein (GP) of filoviruses is indispensable for recognition of its cellular receptor and infection of target cells. To study the intracellular trafficking of GP by using live-cell imaging, the mucin-like domain of Marburg virus (MARV) GP was replaced by the fluorophore mCherry (GP∆MLD_mCherry). Intracellular distribution, surface transport, and recruitment of GP∆MLD_mCherry into virus-like particles were similar to observations for wild-type GP. Using reverse genetics, we generated a recombinant MARV expressing GP∆MLD_mCherry (recMARV MARVGP∆MLD_mCherry). Time-lapse microscopy of recMARV MARVGP∆MLD_mCherry-infected cells revealed that GP∆MLD_mCherry-positive vesicles were transported to the cell surface in a tubulin-dependent manner. Moreover, dual-color live-cell imaging revealed cotransport of GPΔMLD_mCherry and VP40 and their colocalization at the plasma membrane. In this proof-of-concept study we showed that the newly developed GP∆MLD_mCherry is a promising tool to elucidate intracellular trafficking and assembly pathways of MARV.