Identifying Cassia Seed Decoction Pieces Based on the ITS2 Sequence
Objective To distinguish cassia seeds and their counterfeits,and provide a scientific basis for the identification of the origin of cassia seeds.Methods The DNA of cassia samples was extracted by CTAB,PCR amplified,electrophoresed and sequenced;sequences of their counterfeits were downloaded from GenBank.The ITS2 sequence was obtained based on the Hidden Markov Model annotation.MEGA6.0 software was used to analyze the sequence features,calculate the intraspecific and interspecific genetic distances,and construct a phylogenetic tree;Prediction of secondary structure of different basal origin cassia seeds and their counterfeits using ITS2 database.Results Multiple stable SNP sites were found in the ITS2 sequence,which could be used to identify different origin of cassia seeds and their counterfeits based on single or multiple SNP sites.The cassia seeds from different origin had different ITS2 secondary structures and were clustered into a single unit in the phylogenetic tree with good monophyly,which could distinguish cassia seeds with different basal origin and their counterfeits in a more intuitive way.Five of the available cassia seed decoction pieces were identified as Cassia tora,and the rest were Cassia obtusifolia,and no counterfeit products were found.Conclusion The ITS2 sequence can effectively distinguish cassia seeds of different origins and their counterfeits.
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