As a typical representative of tourism resources, the spatial distribution of A-level scenic spots has a profound impact on the layout of tourism industry. Scenic spot accessibility is also important for the development of tourism. However, the relationship of regional accessibility and spatial distribution of A-level scenic spots are understudied. The study used quantitative geography and geographic information system spatial analysis methods and analyzed the evolution of spatial distribution and regional accessibility of A-level scenic spots in Guangdong Province from 2001 to 2020. The results present the following: 1. Agglomeration distribution is the main distribution type of A-level scenic spots in Guangdong Province, and the spatial distribution is unbalanced. 2. From 2001 to 2020, the spatial distribution of A-level scenic spots in 21 prefecture-level cities of Guangdong Province has gradually developed from "wide gap" to "relatively reasonable." 3. Distribution density of A-level scenic spots in Guangdong Province has evolved into the main core area of high density. 4. Center of the gravity of A-level scenic spots in Guangdong Province developed from east to west during 2002–2007 and moved to the east after 2007. 5. Accessibility between A-level scenic spots and tourist source areas in Guangdong Province is good, with an evident aggregation phenomenon. This study reveals the spatial distribution evolution law and regional accessibility of A-level scenic spots, which is conducive to healthy, sustainable, and stable development of tourism in Guangdong Province.