In order to understand sexual reproductive characteristics of an arsenic hyperaccumulator, Pteris vittata L., in soil culture and provide valuable suggestions for its commercial production, two experiments were performed focused on how storage time of spores, collection time, substrate sterilization methods, and culture conditions affect sexual reproductive efficiency. Results demonstrate that spores of P. vittata remain at high viability after several month storage, can live through hot water and spores collected in winter have a higher viability than those collected in spring. Furthermore, light is indispensable for gametophyte growth of P. vittata. During the gametophyte phase and the early sporophyte phase, P. vittata demonstrated a wide adaptability to differing light intensities, but grew best under 40% shade. Additionally, it grew in sand though it prefers fertile substrates. A proper combination of light, substrate, and densities of spores and gametophytes promises efficient reproduction.