In this paper, a research is conducted into vertical vibration problem for large-span floor based on the construction of the long-span prestressed secondary beam roof in the project of a primary school gymnasium, with the structural arrangement of secondary beam height, floor thickness, and the thickness of the flat beam floor changed. Considering the possible load conditions, the natural vibration frequency and peak acceleration of the floor under three load conditions are simulated by ANSYS. After comparing and analyzing the results, the main conclusions are as follow. The increase of floor thickness has little or even negative effect on the vibration characteristics and vibration acceleration of floor; the secondary beam height increases by 20% (1 000~1 200 mm) and 25% (1 200~1 500 mm), the floor frequency of the floor increases by 10.4% and 12.5%, and the decrease rate of RMS acceleration of non-resonance is 20.34% to 36.14%. After adding 50mm thick flat beam floor, the floor frequency of the floor increases by 17.6% when compared with no floor, and the RMS acceleration decrement is 60.67%for all conditions during non-resonance; however, the effect of changing the floor thickness of flat beam on the natural frequency and vibration acceleration of the floor is minimal; the best effect of adding flat beam floor slab to improve floor vibration comfort is to synthesize each working condition.