MODIS-EVI, climate factors and meteorological drought index SPEI were used as the indicators to characterize the spatio-temporal changes of vegetation and climate, Theil-sen Median trend analysis, Mann-Kendall significance test, Hurst index analysis and residual analysis were used to explore the response of vegetation growth to climate change and human activities in Guizhou Province from 2001 to 2020. The results showed that:(1) The regional area ratio of vegetation increase in Guizhou Province from 2001 to 2020 was 84.97%, and the main improvement areas were mainly in the north and southwest of Guizhou Province. In terms of the future spatial change trend, vegetation was mainly in reverse continuous increasing, accounting for 69.23% of the total area of Guizhou Province. (2) There was a positive correlation between vegetation and precipitation and a negative correlation between vegetation and temperature in Guizhou Province, but the correlation was weak. While the positive correlation between regional vegetation growth and SPEI was greater than the negative correlation. (3) In the northern, western and southwestern regions of Guizhou Province, EVI and human activities showed positive improvement, while in Guiyang, Zunyi and its surrounding areas, EVI and human economic activities showed inhibitory effect. On the whole, there was a positive correlation between human economic activities and vegetation EVI in the study area. The conclusion could provide objective scientific theoretical support for the study of vegetation ecological protection and its suitability with climate change and human activities in Guizhou Province.