Innovation Project of Guangxi Graduate Education
Objective To investigate DNA barcode(s) most appropriate for distinguishing Stephania tetrandra from other species. Methods DNA was extracted from fresh leaves of 32 individuals of S. tetrandra and 1 individual of S. excentrica by using the centrifugal column type plant genome kit. The PCR products were amplified with universal primers for the ribosomal DNA second internal transcribed spacer (ITS2), chloroplast ribulose-1,5-bisphosphate carboxylase large subunit (rbcL), and chloroplast non-coding region (psbA-trnH) sequences, followed by electrophoresis and sequencing. DNAMAN software was used for post-sequencing sequence proofreading and assembly, and MEGA7.0 software was used for sequence variation analysis. Moreover, barcode sequences of S. excentrica and 3 different species were retrieved from the GenBank. Neighbor (NJ) joining method was used to construct the phylogenetic tree, and RNAFOLD was used to predict the secondary structure of the sequences.