Abstract:Evodia rutaecarpa(Juss.) Benth is an important medicinal plant species that has diffierent varieties, namely E.rutaecarpa(Juss.) Benth. var. officinalis(Dode) Huang, and E.rutaecarpa(Juss.) Benth. var. bodinieri(Dode) Huang. AFLP technique and ITS region sequencing were used to distinguish 14 samples belonging to the three varieties of E.rutaecarpa(Juss.) Benth. Sequence analysis of the ITS regions amplified from the three varieties of E.rutaecarpa(Juss.) Benth. indicated that ITS regions were highly similar and the identities was more than 97%. A SNP nucleotide was discovered between E.rutaecarpa(Juss.) Benth. and its two varieties. Amplified fragment length polymorphism was compared among E.rutaecarpa(Juss.) Benth, E.rutaecarpa(Juss.) Benth. var. officinalis(Dode) Huang and E.rutaecarpa(Juss.) Benth. var. bodinieri(Dode) Huang and different specific bands appeared among three varieties of E.rutaecarpa(Juss.) Benth. Based on these specific markers, molecular authentication methods can be developed to identify and distinguish E.rutaecarpa(Juss.) Benth and its two varieties.