It is of great value to study the spatio-temporal evolution of construction land with its driving forces in urban agglomeration for the promotion of a high-quality development of territorial space. Taking urban agglomeration in the middle reaches of the Yangtze River as an example, an analysis has been made of the spatio-temporal characteristics and driving mechanism of the evolution of urban agglomeration construction land by using multi-stage Landsat TM/ETM data, expansion intensity index, multiple regression and geographic detector. The results show that from 2000 to 2020, the construction land of urban agglomeration in the middle reaches of the Yangtze River shows a continual expansion trend, with a diversity in the growth area and expansion intensity in different periods. Throughout the study period, the number of urban-dominated, moderately-developed and lowly-developed patches has increased significantly, while the number of natural ecological patches has decreased significantly. There is a significant diversity in the intensity of construction land expansion among different regions within the urban agglomeration, presenting an overall spatial pattern of “polycentric diffusion”. Population density, GDP density and distance from traffic lines are the main driving forces for the expansion and evolution of construction land in urban agglomeration, with the influence of multiple factor interaction greater than that of a single factor. Therefore, it is suggested to strengthen the radiation and driving role of central cities, seeking the promotion of the industrial development and supporting construction of small and medium-sized cities, establishment of mechanisms and paths for an effective cooperation between large and medium-sized cities, facilitation of the reform and improvement of land systems and policies, activation of the urban stock land, and acceleration of the high-quality development of land and space of urban agglomeration in the middle reaches of the Yangtze River.