Differential absorption lidar (DIAL) is an excellent technology for atmospheric CO2 detection. However, the accuracy and stability of a transmitted on-line wavelength are strictly required in a DIAL system. The fluctuation of a tunable pulsed laser system is relatively more serious than that of other laser sources, and this condition leads to a large measurement error for the lidar signal. These concerns pose a significant challenge in on-line wavelength calibration. This study proposes an alternative method based on wavelet modulus maxima for the accurate on-line wavelength calibration of a pulsed laser. Because of the different propagation characteristics of the wavelet transform modulus maxima between signal and noise, the singularities of a signal can be obtained by detection of the local modulus maxima in the wavelet transform maximum at fine scales. Simulated analysis shows that the method is more accurate than the general method such as quintic polynomial fitting and can steadily maintain high calibration precision at different signal-to-noise ratios (SNRs). Last, 16 groups of real experiments were conducted to verify the simulated analysis, which shows that the proposed method is an alternative for accurately calibrating an on-line wavelength. In addition, the proposed method is able to suppress noises in the process of wavelength calibration, which gives it an advantage in accurate on-line wavelength calibration with a low SNR.Research Funds for the Central Universities (2042015kf0015).