ABSTRACT
OBJECTIVE: To evaluate the accuracy and stability of ITS/ITS2 barcodes in identification of Zanthoxmli Pericarpium. METHODS: Total genomic DNAs from samples were extracted by using improved DNA extraction kits. ITS regions were amplified, and the ITS2 sequences were obtained by using the hidden Markov model (HMM) based on annotation method from the ITS sequences. The inter- and intra-specific variation of the Zanthoxmli Pericarpium and its adulterants were analyzed. Zanthoxmli Pericarpium was i-dentified through the Species identification System for Traditional Chinese Medicine and neighbor-joining (NJ) phylogenetic trees. RESULTS: The lengths of ITS/ITS2 sequence of Zanthoxmli Pericarpium were 618-620 bp and 224 bp. The intra-specific genetic distances were smaller than inter-specific ones in ITS/ITS2 regions of Z. schinifolium and Z. bungeanum. Zanthoxmli Pericarpium and its adulterants could be easily differentiated according to the nearest distance, the Species Identification System for Traditional Chinese Medicine and the NJ trees methods. CONCLUSION: ITS/ITS2 regions as DNA barcodes can stably and accurately distinguish Zanthoxmli Pericarpium and its adulterants.
ABSTRACT
In this research, Casuarina eguisetifolia Linn was used to verify the broadly suitability of DNA bar-codes in identification of Li-medicine plants and systematic development of species. The genomic DNA of 22 samples collected C. eguisetifolia and its adulterants were amplified by 4 pairs of primers respectively (ITS (inter-nal transcribed spacer), ITS2 (internal transcribed spacer 2), trnH-psbA , rbcL) and sequenced bi-directionally. Obtained sequences were assembled using CodonCode Aligner. The dates were analysised using MEGA5.1 in ac-cordance with the kimura 2-parameter (K2P) model. The neighbor-joining (NJ) phylogenetic trees were construct-ed. Our study demonstrated the efficacy of ITS/ITS2 to distinguish between C. eguisetifolia and other adulterants species at the molecular level. Comparative to the primer of trnH-psbA and rbcL, there was a obviously DNA gap. The NJ trees showed that the several species of Casuarina can be classified to same types to show a obvi-ously monophyly, which the nearest family was Guttiferae. Therefore, ITS/ITS2 regions can accurately distinguish the original plant of Li-medicine. The systematic evolution of Casuarina can be verified in the molecular level.
ABSTRACT
The ITS/ITS2 barcodes were used to simply and effectively identify Codonopsis Radix and its adulter-ants. In this study, ITS (internal transcribed spacer of unclear ribosomal DNA) regions were amplified using PCR (polymerase chain reaction) from thirty-three samples of Codonopsis Radix and ITS2 regions were obtained from the ITS sequences using the hidden Markov model (HMMer)-based annotation methods. The sequences of ITS/ITS2 regions were aligned and the genetic distances were computed by MEGA5.0. Species identification efficiency of ITS/ITS2 sequences were evaluated using BLAST1 and nearest distance methods. The results indicated that The sequences lengths of ITS regions of Codonopsis Radix were 654-655 bp, and the lengths of ITS2 regions were 239 bp. The intraspecific genetic distances among Codonopsis Radix were obviously lower than the interspecific genetic distance between Codonopsis Radix and its adulterants. Therefore, ITS/ITS2 regions can stably and accu-rately distinguish Codonopsis Radix and its adulterants.