The invention discloses a method for estimating a Doppler spread factor of a MIMO-OFDM underwater acoustic communication system by using a zero correlation zone (ZCZ) sequence as a training sequence. The method comprises the following steps: inserting two segments of repeated ZCZ sequences in front of a data frame, carrying out delayed autocorrelation operation on received signals by a receiving end by adopting a plurality of parallel correlators, and estimating the Doppler spread factor according to the window length of the correlator having the maximum output result. Compared with the existing Doppler spread estimation algorithm, the method can significantly improve the Doppler spread estimation accuracy and has strong channel adaptability, when the signal to noise ratio is greater than -5dB, the estimation accuracy is always kept to be 10<-4> or below, and the requirements of underwater acoustic communication with a high data rate for the estimation accuracy of the Doppler spread factor are satisfied.