Brachiaria brizantha is a C4 grass commonly used as forage in the tropics. Leaf samplesrnfrom seven populations originating from Welega, Jimma, Omo, Gondar, Gojam, Borenarnand Ilu Ababora were collected from Zeway Regeneration and Conservation Site of ILRI,rnEthiopia. Genomic DNA was extracted from 79 accessions each from three individualsrnusing CTAB extraction method. A diluted genomic DNA was subjected to PCRrnamplification. From the ISSR primers tested, only six amplified 80 scorable bands fromrnthe 79 accessions used. 96.25% Percentage of polymorphic loci (PPL), gene diversity (h)rn= 0.366 and Shannon informationâ€™s index (I) = 0.539 were detected at species level. Thernnumber of bands identified by each primer ranged from 8 to 19, with an average of 13.3.rnThe highest genetic diversity was generated from accessions collected from Ilu Ababorarn(PPL= 58.75 %, h=0.210 and I =0.315) while the least was from those collected fromrnGondar region (PPL=28.75%, h=0.124 and I =0.183). Jaccardâ€™s similarity coefficientsrnranged from 0.302 to 0.431 and analysis of molecular variance indicated the presence ofrnhigher proportion of variation within population (64.66%) than between populationsrn(35.34 %). Cluster analysis using the un-weighted paired group method with arithmeticrnaverage (UPGMA) and Jaccardâ€™s similarity coefficient (0.38) clustered the accessionsrninto three major (I, II and III) clusters in their respective regional collection. PrincipalrnComponent Analysis (PCO) showed accessions in populations formed their own cluster.rnThus ISSR markers detected a range of genetic diversity from Brachiaria brizantharngermplasm collections from Ethiopia.