Simple and effective methods are needed for the identification of Chinese medicinal material species and their variety. Lonicera japonica Thunb. is one of Chinese herbal medicines widely demanded. A total of 3 705 EST-SSRs of L. japonica and 2 818 EST-SSRs of L. japonica var. chinensis Thunb. were identified from EST database in our lab. In average, there was one EST-SSR per 4.05 kb in L. japonica ESTs and per 7.49 kb in L. japonica var. chinensis ESTs, separately. The identified SSRs in L. japonica were consisted of 51.98% dinucleotide and 34.61% trinucleotide repeats, while SSRs in L. japonica var. chinensis had 57.45% dinucleotide and 30.09% trinucleotide. The results reviewed that the classes AG/TC and GAG/TCT were predominant in the dinucleotide motifs and the trinucleotide motifs, respectively. Total 87 EST-SSRs were identified of significant difference between L. japonica and L. japonica var. chinensis. PCR products were obtained from 52 L. japonica samples in 13 out of 15 SSR markers tested. The polymorphism in L. japonica, L. japonica var. chinensis and other honeysuckles could be distinguished by three markers (jp.ssr4, jp.ssr64 and jp.ssr65) tested.