木香、川木香、土木香、青木香和红木香药材的ITS2条形码分子鉴定

通过分析木香、川木香、土木香、青木香、红木香药材的ITS2（internal transcribed spacer 2）条形码序列，探讨木香类药材鉴定新方法。对60份样品提取基因组DNA，PCR扩增ITS2序列并进行双向测序，所得序列经CodonCode Aligner拼接后，采用 MEGA5.0 软件进行序列比对，计算种内和种间遗传距离（K2P），构建邻接树（neighbor-joing tree，NJ Tree）。结果表明，无论是菊科的木香、川木香、土木香，还是马兜铃科的青木香和五味子科的红木香，ITS2序列种内变异均小于种间变异，ITS2序列所有单倍型比对后长度为272 bp，变异位点达到162个；遗传距离显示物种种内平均K2P远远小于种间平均K2P；NJ树结果显示木香、川木香、土木香、青木香和红木香药材可明显区分，且能鉴别川木香的2个基原物种。因此，ITS2序列可用于鉴定木香、川木香、土木香、青木香、红木香药材，为临床安全用药提供依据。

Molecular identification of Aucklandiae Radix, Vladimiriae Radix, Inulae Radix, Aristolochiae Radix and Kadsurae Radix using ITS2 barcode

In order to identify Aucklandiae Radix, Vladimiriae Radix,Inulae Radix,Aristolochiae Radix and Kadsurae Radix using ITS2 barcodes, genomic DNA from sixty samples was extracted and the ITS2 (internal transcribed spacer) regions were amplified and sequenced. The genetic distances were computed using MEGA 5.0 in accordance with the kimura 2-parameter (K2P) model and the neighbor-joining (NJ) phylogenetic tree was constructed. The results indicated that for Aucklandiae Radix (Aucklandia lappa), Vladimiriae Radix (Vladimiria souliei and V. souliei var. cinerea),Inulae Radix (Inula helenium),Aristolochiae Radix (Aristolochia debilis) and Kadsurae Radix (Kadsura longipedunculata), the intra-specific variation was smaller than inter-specific one. There are 162 variable sites among 272 bp after alignment of all ITS2 sequence haplotypes. For each species, the intra-specific genetic distances were also smaller than inter-specific one. Furthermore, the NJ tree strongly supported that Aucklandiae Radix, Vladimiriae Radix,Inulae Radix,Aristolochiae Radix and Kadsurae Radix can be differentiated. At the same time, V. souliei (Dolomiaea souliei) and V. souliei var. cinerea(D. souliei var. cinerea) belonging to Vladimiriae Radix were clearly identified. In conclusion, ITS2 barcode could be used to identify Aucklandiae Radix, Vladimiriae Radix,Inulae Radix,Aristolochiae Radix and Kadsurae Radix. Our study may provide a scientific foundation for clinical safe use of the traditional Chinese medicines.