2.2. DNA extraction and PCR amplification
The DNA of herbal materials was extracted with a plant genomic DNA extraction kit (Tiangen Biochemical Technology (Beijing) Co., Ltd, China), and partially optimized regarding the DNA barcoding principles for traditional Chinese herbal medicine (Chen et al., 2014a) and previous research (Liu et al., 2017). Then four regions, ITS2, psbA-trnH , matK, and rbcL , were selected for DNA barcodes. Their primers and amplification conditions were set in accordance with the previous study (Chen et al., 2010). PCR products were confirmed by gel electrophoresis on 1.0% agarose gels after staining with GelRed nucleic acid gel stain (Biotium, USA).
The sample pretreatment and meta-genomic DNA extraction of the mock and pharmaceutical samples were performed following the previously published related research protocols (Liu et al., 2021a).100 mg of each sample was collected for sample pretreatment. DNA concentration was evaluated using spectrophotometric measurement of absorbance at 260 nm wavelength (NanoDrop one, Thermo Fisher Scientific Inc., USA).