Specific PCR Identification between Peucedanum praeruptorum and Angelica decursiva and Identification between Them and Adulterant Using DNA Barcode.

Abstract

Background:The traditional Chinese medicine (TCM) Qianhu and Zihuaqianhu are the dried roots of Peucedanum praeruptorum and Angelica decursiva, respectively. Since the plant sources of Qianhu and Zihuaqianhu are more complex, the chemical compositions of P. praeruptorum and A. decursiva are significantly different, and many adulterants exist because of the differences in traditional understanding and medication habits. Therefore, the rapid and accurate identification methods are required.Objective:The aim was to study the feasibility of using DNA barcoding to distinguish between Traditional Chinese medicine Qianhu (Peucedanum praeruptorum), Zihuaqianhu (Angelica decursiva), and common adulterants, based on internal transcribed spacer (ITS) sequences, as well as specific PCR identification between P. praeruptorum and A. decursiva.Materials and Methods:The ITS sequences of P. praeruptorum, A. decursiva, and adulterant were studied, and a phylogenetic tree was constructed. Based on the ITS barcode, the specific PCR primer pairs QH-CP19s/QH-CP19a and ZHQH-CP3s/ZHQH-CP3a were designed for P. praeruptorum and A. decursiva, respectively. The amplification conditions were optimized, and specific PCR products were obtained.Results:The results showed that the phylogenetic trees constructed using the BI and MP methods were consistent, and P. praeruptorum and A. decursiva sequence haplotypes formed their own monophyly. The experimental results showed that in PCR products, the target bands appeared in the genuine drug and not in the adulterant, which suggests the high specificity of the two primer pairs.Conclusion:The ITS sequence was ideal DNA barcode to identify P. praeruptorum, A. decursiva, and adulterant. The specific PCR is a quick and effective method to distinguish between P. praeruptorum and A. decursiva.SUMMARY Peucedanum praeruptorum and Angelica decursiva sequence haplotypes formed their own monophyly.The ITS sequence was ideal DNA barcode to identify P. praeruptorum, A. decursiva, and adulterant.Specific PCR is a quick and effective method to distinguish between P. praeruptorum and A. decursiva. Abbreviations used: TCM: The traditional Chinese medicine, P.: Peucedanum, A.: Angelica, ITS: The internal transcribed spacer, PCR: Polymerase chain reaction, NCBI: National Center for Biotechnology Information, NI: Number of individuals, HN: Haplotype number; GAN: Gen Bank accession numbers, L.: Ligusticum, O.: Ostericum, A.: Angelica, P.: Pimpinella, BI: Bayesian inference, MP: Maximum parsimony, AIC: Akaike Information Criterion, MCMC: Markov Chains Monte Carlo, TBR: Tree bisection-reconnection, LPP: Length of PCR product, PRP: PCR reaction procedure, SNP: Single nucleotide polymorphisms, PP: Posterior probability, BS: Bootstrap.Qun Zhao