Germination of platanus acerifolia's winter buds and cones falling bring great inconvenience to people's lives, therefore, how to reduce it becomes an issue of concern. Now studies on platanus acerifolia are mostly from phenotypic variation and follow-up observations which had difficulty in guaranteeing long-term stability, but less at the molecular level. This experiment use young leaves of fine individual none-or-little cones platanus acerifolia as experimental materials, adopt suitable CTAB method to extract DNA, then sixty-five ISSR primer were screened by two different samples, eleven high-definition, repeatability and strong stability ISSR primer combinations were chose for the ISSR analysis of the fourteen platanus acerifolia materials. Do Cluster Analysis with NTSYS software, constructing system trees. The results show that the genetic similar coefficients (GSC) were 0.58–0.77, and 14 materials were divided into 5 groups at GSC 0.67 based on the cluster analysis of UPGMA method.