Abstract: ISSR was used to study the genetic diversity and genetic relationship of 37 Castanea henryi native varieties. 13 ISSR primers were selected from 65 universal primers. The number of bands produced by each primer varied from 9 to 16, the average was 12. A total of 156 bands were produced, in which 129 were polymorphic, the percentage of polymorphic loci was 82.69%, in average 9.9 polymorphic bands produced by each primer,and a relatively high level of genetic diversity was revealed by the result. The Observed number of alleles was 1.826 9, the effective number of alleles was 1.509 7, Nei's gene diversity was 0.292 3, Shannon information index was 0.434 4. 37 cultivars were distinguished by 13 ISSR primers. A dendrogram showed that the genetic relationship was constructed through an unweighted pair group method (UPGMA) based on the genetic identity and 37 cultivars were clustered into 2 main groups and 7 subgroups.