Abstract: AIM To study the genetic diversity of ITS sequences of Herba Dendrobii(Huangcao) and analyze the utility of ITS sequences in molecular authentication of Herba Dendrobii (Huangcao)and phylogenetic of Dendrobium. METHODS The ITS gene fragment was amplified using a pair of primers. The PCR products were purified and sequenced by the methods of Sanger Dideoxy. RESULTS The DNA sequence of 228-233 bp ITS 1, 242-247 bp ITS 2 gene fragments and 5.8S rDNA were obtained from 14 samples of Dendrobium. The interspecific substitution varies from 11.79% to 31.58% at ITS 1 and 10.29% to 25.30% at ITS 2. The intraspecific substitution of D.nobile is 0.87% at ITS 1 and without difference at ITS 2. The substitution between Dendrobium and outgroup varies from 23.56% to 36.89% at ITS 1 and 26.52% to 33.31% at ITS 2. The phylogenetic tree based on ITS 1 and ITS 2 data was set up. CONCLUSION The ITS 1 and ITS 2 gene fragments were highly conservative at intraspecific level in Dendrebium, while they were less conservative at interspecific level in D.nobile. They were least conservative between Dendrobium and outgroup. Hence, the sequence of this fragment is a good molecular marker for authentication of the Huangcao. But, further study is necessary for the phylogenetic of Dendrobium.