Because of the measurement error, the measurement result is uncertain. Measurement uncertainty is an important indicator for evaluating the quality of measurement results. The ratio of the sampling length to the signal period is divided into integer and decimal parts, and the measurement errors of the DFT frequency and phase difference are deduced. The Class B uncertainty evaluation method is adopted to analyze the frequency measurement uncertainty caused by spectrum leakage, windowing, and fence effect, and the synthetic uncertainty of frequency measurement is obtained. Then the phase difference uncertainty is determined through the relationship between phase difference measurement error and frequency measurement error. Finally, through calculation and analysis, the relationship between the sampling length and measurement error as well as uncertainty of frequency and phase difference is verified. The analysis results show that the evaluation process is feasible and effective.