Development of cpDNA markers for discrimination between Cynanchum wilfordii and Cynanchum auriculatum and their application in commercial C-wilfordii food products

Cynanchum wilfordii has been used as a health-enhancing food product in Korea. Owing to its morphological similarity to C. wilfordii and relative ease of cultivation, C. auriculatum has been illegally used as a substitute plant for C. wilfordii. In the present study, we developed markers to discriminate between C. wilfordii and C. auriculatum. Species-specific primer sets for C. wilfordii and C. auriculatum were designed based on the single nucleotide polymorphisms of the chloroplast matK genes for quantitative real-time PCR using SYBR green. The limit of detection of primer sets for each species was assessed by analyzing serially 1/10-diluted DNAs at concentrations of 0.001-100% (=10 ng) and binary mixtures of a flour matrix spiked with decreasing concentrations (10(3)-1 mg/g) of non- and heated C. wilfordii and C. auriculatum, respectively. We found that species-specific primer sets indicated good amplification efficiency and correlation coefficients (R (2)) of the standard curves in the extracted DNA. The developed markers were successfully applied to 19 commercial C. wilfordii food products and could prove a useful tool for verifying the presence of C. wilfordii and C. auriculatum in commercial products.