Development of a species-specific real-time PCR assay for identifying the medicinal plant Artemisia princeps and the toxic plant Corydalis speciosa

Abstract The medicinal herb Artemisia princeps and the poisonous plant Corydalis speciosa have a high degree of morphological similarity, which may contribute to unintentional contamination. Indeed, cases of acute toxicity have been reported in which C. speciosa was mistakenly ingested in place of A. princeps . To ensure accurate identification, a species-specific quantitative real-time PCR (qPCR) assay was developed using six primer pairs targeting single-nucleotide polymorphisms or insertion/deletion regions within chloroplast genes. The respective limits of detection were validated, and a contamination-based cutoff Cq value (0.1%) was established for assessing authenticity. All six primer pairs showed excellent performance, with correlation coefficients > 0.99, slopes of -3.12 to -3.28, and efficiencies of 101.82%–109.47%. The developed qPCR assay was verified based on blind and commercial tests for reproducibility and specificity, respectively. This qPCR assay represents a sensitive and reliable method for distinguishing similar medicinal and toxic plants, thereby contributing to improved product quality and safety.