Medium-Mn steel is the newly developed steel acting as a promising candidate of the 3rd-generation advanced high strength steels. In the present study, the effect of hydrogen on the mechanical behavior and martensite transformation process in a duplex medium-Mn steel is investigated by in-situ nanoindentation test. The mechanical response of individual phase: ferrite, and retained austenite by introducing hydrogen is studied. With the presence of hydrogen, the reduction of activation energy for dislocation nucleation was verified by using a stress-biased, statistical thermal activation model. The stacking fault energy (SFE) was reduced, which was revealed by electron channeling contrast (ECC) technique. Magnetic force microscopy (MFM) revealed a suppression of retained austenite to α′-martensite transformation with the presence of hydrogen, which is related to hydrogen induced SFE reduction and enhanced slip planarity.