Polymorphic expressed sequence tag - simple sequence repeat (EST-SSR) markers derived from major cereal crops were used to assess the genetic diversity of the USDA temperate bamboo collection consisting of 92 accessions classified in 11 separate genera and 44 species. A total of 211 bands were detected with a mean number of alleles per locus of 8.440. Phylogenetic relationships were determined by calculating genetic distances between all pairwise combinations and assessing differences in character data. The resulting dendrograms (unweighted pair group method with arithmetic means (UPGMA) and parsimony) clustered the accessions into 2 main clades, which corresponded to accessions characterized morphologically as either clumping (sympodial) or running (monopodial) bamboos. The majority of the accessions clustered according to their current taxonomic classification. These markers were also beneficial in identifying contaminated and (or) misidentified plots. Overall, these transferred markers were informative in differentiating the various bamboo accessions and determining the level of genetic variation within and among species and genera.