Identification of Bletillae Rhizoma and its adulterants by SNPs in ITS2 / 中国中药杂志

ABSTRACT

To establish a molecular identification method for Bletillae Rhizoma, this paper extracted genome DNA from Bletillae Rhizoma and its adulterants. The sequences of rDNA ITS2 were sequenced after amplifying. Then multiple alignments of ITS2 were constructed phylogenetic tree with Neighbor Joining by MEGA 5. 1 and found out SNPs loci. The result showed that rDNA ITS2 region could identify Bletillae Rhizoma and its adulterants. There existed the SNPs loci, which could identify Bletilla striata and B. ochracea. Furthermore, we designed specific primers against the SNPs loci of B. striata and B. ochracea, then screened primers and optimized the PCR amplification conditions. Finally, the DNA of B. striata and B. ochracea were specifically amplified by BJ59-412F, BJ59-412R and HHBJ-225R. The length of amplification products were respectively about 350 bp and 520 bp that were effectively identified of B. striata and B. ochracea. While, the adulterants of Bletillae Rhizoma were no-reaction occurring. To sum up, the amplification conditions of the primers can identify B. striata, B. ochracea and their adulterants successfully at the same time. This method was easy, time-saving, and reliable, which can be used as a rapid method for molecular identification of Bletillae Rhizoma.