Monitoring and identification of Cynanchum wilfordii and Cynanchum auriculatum by using molecular markers and real-time polymerase chain reaction

Abstract

Cynanchum wilfordii (Asclepiadaceae) is widely distributed throughout Korea, Japan, and China. Dried roots of this plant have been used as a tonic to promote renal function. Due to the morphological similarities of the dried roots of this plant to those of Cynanchum auriculatum, which is used as a substitute herbal medicine for C. wilfordii, distinguishing these two species is extremely difficult. The present study was conducted to develop molecular markers to distinguish C. wilfordii and C. auriculatum by using conventional polymerase chain reaction (PCR) and realtime PCR analyses. Comparative analysis based on the sequence of the trnL-trnF intergenic spacer revealed 4 base-pair variations, and the inter-individual sequences of the 2 species separately showed 100% homology. According to these results, the variations were divided into 2 groups. The 2 species were further distinguished using a sequence-characterized amplified region marker developed based on a randomly amplified polymorphic DNA-PCR product, and then a single nucleotide polymorphism marker was designed based on the trnL-trnF intergenic spacer for more efficient detection in real-time PCR. The results showed that speciesspecific molecular markers might allow accurate discrimination of C. wilfordii and C. auriculatum.