The genetic diversity of the 29 accessions of Pennisetum longissimum var.intermedium from Gansu and Yunnan were analyzed by ISSR makers.The results showed that:(1)10 primers were selected from 50 primers which had clear,stable polymorphic bands.A total of 72 loci were obtained,including 62 polymorphic loci.On average,amplification site of each primer was 7.2 and the percentage of polymorphic loci (PPL) was 87.4%.The mean observed number of alleles (Na),effective number of alleles (Ne),Shannon information index (I) and Nei’s genetic index (H) were 1.861 1,1.742 8,0.561 0 and 0.395 9,respectively.The genetic similarity coefficient (GS) ranged from 0.236 to 0.903,which shows rich genetic diversity among the materials of P.longissimum var.intermedium.(2)Cluster analysis with UPGMA method showed that the 29 tested accessions were divided into 4 groups at the level GS 0.51,but there was no correlation (r=0.437 0,P=0.204 6) between genetic and geographic distance among the germplasms studied.The results of this research were the first time to reveal genetic diversity and variation of P.longissimum var.intermedium at the molecular level,which provided theoretical and data evidence for the introduction,domestication,conservation and exploitation of wild P.longissimum var.intermedium germplasm resources.