Abstract: In order to know pitaya germplasm resources distribution and the genetic relationship in Fujian, an investigation on pitaya planting was conducted, and the samples were also collected. ISSR technology was applied to analyze the genetic diversity of 20 germplasm resources collected. 11 appropriate primers were selected from 40 random ISSR primers, 142 scorable amplification sites were detected, and the percentage of polymorphic ratio was 74.65%. The result of cluster analysis based on Jaccard revealed that the similarity coefficients (GS) between different cultivars ranged from 0.478 to 0.937, the average was 0.646. And when GS was 0.559, the whole population could be divided into 3 groups. This paper preliminarily revealed the pitaya germplasm resources distribution and the genetic relationship in Fujian and also proposed some varieties such as ‘Baijulong’, ‘Luobai’ and ‘Hongxianmi 1’ who suitable for popularization, which provided valuable reference for the species selection and breeding of pitaya.