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The scale research of landscape pattern is an important basis for the study of spatiotemporal evolution of landscape pattern and the scientific and reasonable allocation of landscape pattern. This paper takes the middle and upper reaches of Meijiang river as the research area. Combined with spatial grain size analysis and extent size analysis, the spatial scale effect was studied to select the optimal research scale in this research area. The results show that most of the appropriate grain size of landscape pattern indexes are in range of 90–150 m, and the optimal grain size scale is 150 m in Middle and Upper Reaches of Meijiang River Basin. At the grain size scale of 150 m, the spatial self-correlation of landscape pattern index over 50% is the highest at the extent size of 300 m, and the optimal extent size scale is 300 m in this basin. This paper determines the optimal scale of landscape pattern in the middle and upper reaches of Meijiang river basin, which is of great significance to the ecological balance and sustainable development of the basin.