The Glycyrrhiza L.SSRs were searched in the expressed sequence tag (EST) database of Glycyrrhiza uralensis,and the EST-SSR primers were designed by Primer 3.0 online software.The EST-SSR fingerprint characteristics and cluster trees of four species were then analyzed to discuss the interspecific relationships among species and taxonomic status of the doubtful species.(1)Total 504 SSRs were identified from 441 EST sequences.There were 350 dinucleotide repeat sequences with 69.44% frequency,showing the dominant types in the SSRs.Both TC/AT and TA/AG types were abundant among dinucleotide repeat sequences.(2)The distinct bands were produced in PCR amplifications for all 40 random selected EST-SSR primers.The polymorphic bands were observed in 15 primers,a total of 59 alleles in the 22 materials of the Glycyrrhiza L.were observed,which appeared to be 89.44% polymorphic among these 15 primers with an average of 3.93 alleles per primers.(3)The primer 64 could amplify specific bands in 4 species.G.eurycarpa had the same alleles as that of G.inflata at 180 bp site and that of G.grabra at 220 bp site,showing the hybrid characteristics.(4)The cluster analysis of SSR data showed that when similarity coefficient was 0.82,22 materials from 4 species were clustered into 4 groups:G.uralensis in Hangjingqi,Neimenggu;G.eurycarpa and G.grabra in Bachu,Xinjiang;G.eurycarpa and G.inflata in Shihezi,Xinjiang;G.uralensis in Shihezi,Xinjiang.This cluster was in accord with the classical taxonomy.However,there was fairly large genetic differentiation in different populations of G.eurycarpa.It may be caused by the differentiation of sympatric parent species or introgressive hybridization.These results indicates that 15 primers is suitable for EST-SSR analysis of Glycyrrhiza L..It would be useful for species identification and study of genetic variation and species relationship of Glycyrrhiza L..