Abstract: In this paper, genetic relationship among local cherry cultivar Manaohong originated from Nayong County of Guizhou Province, as well as: sour cherry, black pearl and four sweet cherry cultivars were investigated using ISSR markers. The purpose was to investigate the causes of variation on Manaohong. A total of 286 markers were scored from 15 primers, which were demonstrated highly reproducible, clear and polymorphic bands. Among the obtained markers, 249 were polymorphic, accounting for 87.06% of the total. As analyzed by NTSYS2.01, the genetic similarity among the seven accessions ranged from 0.41 to 0.94, and the genetic similarity between Manaohong and sour cherry was 0.57Manaohong has significant genetic differences with sour cherry on molecular level, possibly it is a hybridization variation of sour cherry.