Establishing DNA markers to differentiate and , which are confusedly used as medicinal herbs, using real-time PCR.

Unlabelled: and are used as medicinal herbs and aromatic plants and belong to the family . Despite differences in composition and physicochemical properties, both plants are frequently sold as the medical substance "Kwakhyang" in some Asian countries. Molecular markers were established to distinguish between the two plants using quantitative real-time PCR. Species-specific primers were designed in the nuclear internal transcribed spacer region of ribosomal DNA and in the chloroplast genes , , and . Six primer sets were tested, the correlation coefficient was higher than 0.99, and the slope was approximately - 3.36 to - 3.58. Efficiency ranged from 90.13 to 98.52%. The developed real-time PCR assay was validated with 14 off-target species, and its reliability was verified through blind testing of 14 commercial products. The assay developed here may help protect consumer rights, and the designed primers can be used to distinguish between the target species.

Supplementary Information: The online version contains supplementary material available at 10.1007/s10068-022-01176-y.