As a new energy-saving air supply mode, stratum ventilation system has not been widely applied due to inadequate theoretical support. A mathematical model based on the characteristics of the indoor air flow was established to predict the vertical temperature profile in a stratum ventilated room. By combining the characteristics of indoor air flow with mass and energy equilibrium equations, this model gave the quantitative relationship between indoor vertical temperatures and boundary conditions such as heat source, enclosure radiation and supply air parameters. A comparative study shows that the predictions of this model fit the experiment values well. The model can represent the practical vertical temperature profiles in a stratum ventilated room. The analyses indicate that this proposed model has acceptable accuracy and can be used for practical engineering designs and energy consumption analysis.