Morphological and Molecular Identification of Original Species of Chebulae Fructus from Adulterants
Objective:To develop a precise molecular identification method for distinguishing Chebulae Fructus(Terminalia chebula),T.chebula var.tomentella,and their adulterants.Method:The fresh fruits of T.chebula,T.chebula var.tomentella,and their adulterants were dried,and their morphological characteristics were observed before and after drying.Sample DNA was extracted,amplified,and sequenced using DNA barcode ITS2 and psbA-trnH sequences via polymerase chain reaction.The sequencing results were aligned using DANMAN 7 and MAGE 6 software to calculate Kimura 2-parameter genetic distance and create a neighbor-joining(NJ)phylogenetic tree.Results:Morphological observations revealed significant differences between the fruits of T.chebula,T.chebula var.tomentella,and their adulterants before and after drying,which could be identified based on morphological distinctions.The DNA barcode sequence showed that the ITS2 sequences of T.chebula and T.chebula var.tomentella were largely similar,with only four single nucleotide polymorphism sites,making them indistinguishable based on this sequence.However,there were two distinct differences in the psbA-trnH sequences between T.chebula and T.chebula var.tomentella,allowing accurate differentiation between these two species.Two haplotypes were identified in T.chebula var.tomentella,with a significant variation related to the presence of a 16 bp tandem repeat sequence in the psbA-trnH sequence.Adulterants could be differentiated from T.chebula,T.chebula var.tomentella based on significant differences in the psbA-trnH sequence and confirmed through the National Center for Biotechnology Information.Identification of processed T.chebula in the market revealed that most samples were two haplotypes of T.chebula var.tomentella,with instances of adulterant T.bellirica.Conclusion:The psbA-trnH sequence can be applied to accurately identifying T.chebula,T.chebula var.tomentella,and their adulterants,discovering two haplotypes in T.chebula var.tomentella.
万方数据
维普
