基于DNA条形码鉴别半夏及其混伪品OA

Objective To establish an accurate identification method for Pinellia ternata and its common adul-terants based on the internal transcribed spacer 2(ITS2)sequence and its secondary structure.Methods 19 bat-ches of samples,including Pinellia ternata,Pinellia pedatisecta,Pinellia cordata,Typhonium flagelliforme,and Ari-saema heterophyllum,were collected.Genomic DNA was extracted,amplified with universal ITS primers,and bidi-rectionally sequenced.11 ITS2 sequences of related species were retrieved from NCBI.Self-generated and database sequences were integrated to calculate intra-and inter-species Kimura 2-parameter(K2P)genetic distances using MEGA 11.0,followed by Neighbor-Joining(NJ)tree construction.ITS2 secondary structures were predicted via online tools.Results Intra-specific genetic distances ranged from 0-0.011,while inter-specific distances spanned 0.055-0.339,demonstrating distinct barcoding gaps.Within the studied taxa,intra-generic genetic distances consis-tently measured below 0.150,whereas inter-generic distances uniformly exceeded 0.150,supporting 0.150 as an em-pirical genus-level threshold.The NJ tree revealed monophyletic clustering for all species except Typhonium gigan-teum.and Typhonium flagelliforme,which failed to form a cohesive clade,and Arisaema heterophyllum,which branched independently,indicating taxonomic inconsistencies.ITS2 secondary structures exhibited a conserved"central loop with four helical arms"across species except Arisaema heterophyllum,with genus-specific variations in loop morphology and helix arrangements enabling reliable differentiation of closely related taxa.Conclusion ITS2 sequence coupled with secondary structure analysis provides a robust tool for authenticating Pinellia ternata and its adulterants while exposing phylogenetic incongruences in traditional classification.This approach is recommended as a complementary DNA barcode for quality control of Pinellia-based medicinal products.