Abstract:The ITS2，psbA-trnH，and matK sequences were used to identify Illicium verum and its adulterants.The total DNA was extracted from Illicium verum，the original plant of Illicium verum，for PCR amplification and sequencing. In addition，80 sequences of 8 species of adulterants were downloaded from GenBank.The standard ITS2 sequence was obtained by Hidden Markov Model（HMM）annotation. According to the annotations of the same species in GenBank，standard psbA-trnH and matK sequences were obtained. MEGA was used to calculate the G，C content，sequence lengths，and intraspecific and interspecific genetic distances and build the neighbor-joining（NJ）tree.The secondary structure of ITS2 was predicted by ITS2 Database and 4Sale was used to align the sequences of both primary and secondary structures. Finally，ProfDist was used to build a profile neighbor-joining（PNJ）tree based on genetic distances. The results showed that the interspecific genetic distance between Illicium verum and its adulterants was significantly longer than the intraspecific genetic distance. The phylogenetic tree showed that Illicium verum branched independently. The ITS2 secondary structure varied between species. The PNJ tree had higher resolution than the NJ tree. Although ITS2，psbA-trnH，and matK sequences can accurately distinguish Illicium verum from its adulterants，the matK sequence did not present an advantage in distinguishing species of the same genus. Therefore，the DNA barcoding based mainly on ITS2 and psbA-trnH and supplemented by matK was recommended for the identification of Illicium verum.