## 1 Introduction
Therefore, we developed additional simple and reliable molecular genetic tools for detecting the adulterants and contaminants of Adenophorae Radix. DNA barcoding is a reliable genetic tool used to identify species and is frequently employed to study phylogenetic relationships [12; 13; 14]. This method also has been used to identify the botanical origins of herbal medicines and for discriminating between authentic product and inauthentic adulterants [13; 15; 16]. Over 17 DNA barcode regions can be used to identify the species of Plantae and to differentiate authentic herbal medicines [1], with the genes RuBisCO (_rbc_L) and maturase-K (_mat_K) in the chloroplast genome being commonly used [17; 18]. The internal transcribed spacer (ITS) sequences within the nuclear ribosomal RNA genes (nrDNA) may be used to distinguish plants at the species level [19]. However, the nrDNA-ITS regions have been of limited use because the available primers for these sequences amplify DNA from fungi, other endophytic microorganisms, and multiple copies of the ITS regions. Recently, ITS2 has been proposed as a universal DNA barcode for identifying plant species, herbal medicines, and herbaceous dietary supplements, but it has some disadvantages, such as the complexity of the experiment, the difficulty of interpreting the results, the limitation of sample numbers, and the detection of contaminants [16; 20; 21]. A sequence-characterized amplified region (SCAR) marker assay can reliably and reproducibly distinguish species based on sequence information obtained from DNA fingerprinting or barcoding in diverse plants and herbal medicines. This method amplifies only target-containing samples using specific primers and differentiates positive or negative amplification of target regions, as well as length polymorphisms of target regions by gel electrophoresis of closely related samples [22; 23; 24]. Therefore, the SCAR method is simple to carry out, its results are simple to interpret, and it produces fewer errors in PCR amplification and sequencing [21]. Furthermore, the efficiency and ability of this assay improved with the combination of individual SCAR markers and multiplex PCR, namely, multiplexed SCAR marker assays [22; 25; 26]. These have been abundantly and successfully applied to authenticate medicinal plant species and commercially-processed herbal medicines [22]. A genetic assay has already been developed to discriminate between authentic Adenophorae Radix species based on 28S and 5S rDNA intergenic spacer region sequences of _A. stricta_, _A. triphylla_, _A. hunanensis_, and _G. littoralis_[4]. Another molecular genetic tool can identify authentic Glehniae Radix from common herbal adulterants, including _A. triphylla_ and _A. stricta_, using ITS2 DNA barcoding [3]. However, there are differences in the range of plant species that adulterate Adenophorae Radix between Korea and China. Therefore, an additional marker is needed to authenticate Adenophorae Radix and to simplify analysis. In this study, we analyzed the entire ITS sequence of six medicinal plant species, _A. stricta_, _A. triphylla_, _A. triphylla_ var. _japonica_, _C. lanccolata_, _C. pilosula_, and _G. littoralis_, and developed SCAR markers to differentiate the four herbal medicines using the ITS1 region. In addition, we established a multiplexed SCAR assay for the simultaneous authentication of these herbal medicines in a single PCR reaction. Furthermore, we confirmed that commercial herbal medicines could be successfully used in the assay. Therefore, our method could be used to reliably detect adulteration and to standardize the quality of Adenophorae Radix.