Abstract: The construction of large-scale water conservancy projects in the upper and middle reaches of the Changjiang River has brought a relatively large impact on the runoff regime in the middle and lower reaches of the Changjiang River. In view of the unreasonable and inconsistent test results in the existing runoff regime research by using single alteration analysis method, the hydrological alteration diagnosis system was used to identify the alteration and divide the alteration period of measured runoff series at Yichang, Hankou and Datong hydrological stations, to conclude the evolution characteristics of the runoff regime in the middle and lower reaches of the Changjiang River. And the attribution analysis method based on rainfall runoff curves was used to study the main influencing factors from the perspective of climate change and human activities. The analysis results showed that there was alteration with different types in 9 time series at Yichang Station, which was the most affected station among all research stations in terms of alteration degree and quantities. From upstream to downstream, the influence degree of Hankou and Datong Stations decreased gradually. The runoff in the flood season of the three stations showed a downward trend, while the runoff in the dry season showed an upward trend except the time series of May at Hankou Station. Attribution analysis showed that human activities were the main cause of hydrological alteration of runoff series in three stations except July at Yichang Station.