Identification of Dictamnus dasycarpus Based on psbK-psbI Sequence
Objective:To identify Dictamnus dasycarpus from 22 batches of samples from Liaoning province and 2 batches of samples from Guizhou province by DNA barcoding based on the psbK-psbI sequence.Methods:The DNA extraction reagent kit was used to extract DNA from the root bark of 24 D.dasycarpus samples.The psbK-psbI sequence was amplified by polymerase chain reaction(PCR)and then subjected to paired-end sequencing.The sequences of Citrus medica and C.sinensis were downloaded from the National Center of Biotechnology Information(NCBI)as the out-groups.SeqMan 7.1.0 was used to assemble the sequencing results,and DNAsp 5.1 was used to calculate the haplotype number(h)and haplotype diversity(Hd)of the sequences.MEGA 7.0 was used to align the sequences.The Kimura 2-parameter(K2P)model was employed to calculate the genetic distance,on the basis of which the phylogenetic tree was established with the neighbor-joining(NJ)method.Results:Among the psbK-psbI sequences of 24 D.dasycarpus samples from Liaoning and Guizhou,21 sequences showed no variation,while 3 sequences had 1-3 mutation sites.The genetic distance and phylogenetic tree showed that all samples of D.dasycarpus were clustered into a large clade,and the out-groups were clustered into the other clade,with the support rate of 100%,which indicated small intra-species differences.Conclusion:The DNA barcoding based on psbK-psbI sequence can distinguish D.dasycarpus from its homologous plants,which proves the genetic stability of D.dasycarpus.This method is stable and can be used for distinguishing between D.dasycarpu and other species,which provides a theoretical basis for the subsequent research on this plant at the molecular level.
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