Identification of Dendrobium officinale Using DNA Barcoding Method Combined with HRM and qPCR Technology

Dendrobium officinale is widely used as a traditional medicine across East Asia. It has several medicinal and ornamental values. Therefore, D. officinale is frequently adulterated with other Dendrobium species in the market. Morphological similarities in Dendrobium species limit the easy identification of individual specimens. In this study, DNA barcoding was used as the sequence model system, and SNP-specific sites were explored for the identification of D. officinale. In this study, a total of 45 Dendrobium species including 25 D. officinale and 20 different Dendrobium species were explored. The identification abilities of 9 DNA barcodes were determined using 45 Dendrobium materials, and the findings showed that they had different identification capabilities. High-resolution melting curve analysis (HRM) technology was used to differentiate D. officinale species from non-D. officinale species, using DNA barcode ITS2 which had the strongest identification capability. Intraspecific sequence variability among 25 D. officinale samples was significantly diverse. DNA barcode sequences analysis identified single nucleotide polymorphism (SNP) sites in 4 DNA barcode sequences (ITS2, ycf1b, psbK-psbI, and psbA-trnH). SNP site-specific primers were designed using the amplification refractory mutation system (ARMS), and intraspecific identification of D. officinale was verified using real-time PCR. In addition, D. officinale samples No. 05 and 24 were identified. The findings of this study provide information on the accurate identification of D. officinale samples. Furthermore, it provides a novel insight into the identification of species with similar evolutionary histories and into determining the quality of medicinal materials.