Systematic conservation planning, a widely used approach to identify priority lands and waters, uses efficient, defensible, and transparent methods aimed at conserving biodiversity and ecological systems. Limited financial resources and competing land uses can be major impediments to conservation; therefore, participation of diverse stakeholders in the planning process is advantageous to help address broad-scale threats and challenges of the 21st century. Although a broad extent is needed to identify core areas and corridors for fish and wildlife populations, a fine-scale resolution is needed to manage for multiple, interconnected ecosystems. Here, we developed a conservation plan using a systematic approach to promote landscape-level conservation within the extent of the South Atlantic Landscape Conservation Cooperative. Our objective was to identify the highest-ranked 30% of lands and waters within the South Atlantic deemed necessary to conserve ecological and cultural integrity for the 10 primary ecosystems of the southeastern United States. These environments varied from terrestrial, freshwater aquatic, and marine. The planning process was driven by indicators of ecosystem integrity at a 4-ha resolution. We used the program Zonation and 28 indicators to optimize the identification of lands and waters to meet the stated objective. A novel part of our study was the prioritization of multiple ecosystems, and we discuss the advantages and disadvantages of this approach. The evaluation of indicator representation within prioritizations was a useful method to show where improvements could be made; some indicators dictated hotspots, some had a limited extent and were well represented, and others had a limited effect. Overall, we demonstrate that a broad-scale (408,276 km² of terrestrial and 411,239 km² of marine environments) conservation plan can be realized at a fine-scale resolution, which will allow implementation of the regional plan at a local level relevant to decision making.