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Alternaria alternata  apple pathotype (previously  A. mali ) causes alternaria blotch disease of apple ( Malus  × domestica ), which may result in leaf spots and up to 70% premature leaf drop in serious cases. This disease is of worldwide importance but is most serious in eastern Asia (Japan, Korea, and China) and in parts of the United States. The excessive use of fungicides not only adds cost to apple growers, but also pollutes the environment. In this study, we characterized a 5-year F 1  population from a cross of a resistant cultivar (Huacui) and a susceptible cultivar (Golden Delicious) consisting of 110 individuals along with 14-year-old parent trees (10 each). A field evaluation of disease severity was conducted in 2008 and 2009 under the natural conditions in Liaoning, China (lat. 40°37′ N, long. 120°44′ E). Based on the field data, 110 F 1  plants were divided into five groups. Artificial inoculation was carried out both on the living trees and on the detached leaves in 2009 to ensure that  A. alternata  apple pathotype was the causative agent. Eighty primer pairs of simple sequence repeat (SSR) were screened against the four genomic DNA pools, respectively, from six highly susceptible F 1  plants, six most resistant F 1  plants, one tree of the seed parent, and the one tree of the pollen parent. One pair of primers (CH05g07) was shown to be linked to the DNA pools of susceptible F 1  and the parent tree, but not to the DNA pools of resistant F 1  and parent trees. This primer pair was then used to screen all individual 110 F 1  progenies and two parent trees. The differentiation of 103 individuals (97.3%) with the marker matched the field disease resistance rating. This marker was further screened with 20 cultivars with known susceptibility or resistance to  A. alternata  apple pathotype and its linkage to susceptibility was validated. These results suggest that this marker can be used in marker-assisted selection for resistance/susceptibility to alternaria blotch disease in apple.