Molecular Identification of Bupleurum chinense Seeds Based on DNA Barcoding Technology
10.13422/j.cnki.syfjx.20200650
- VernacularTitle:基于DNA条形码技术的北柴胡种子分子鉴定
- Author:
Qing ZHAO
1
;
Hong-bo XIE
1
;
La YANG
2
;
Miao-jie WEI
3
;
Lin-chun SHI
3
;
Jin-xin LIU
1
;
Chun-ying ZHAO
1
Author Information
1. Hebei Key Laboratory of Research and Development of Chinese Medicine, Chengde Medical University, Chengde 067000, China
2. School of Medicine, Tibet University, Lhasa 850000, China
3. Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100193, China
- Publication Type:Research Article
- Keywords:
Bupleurum chinense;
seeds;
DNA barcoding;
ribosomal DNA internal transcribed spacer (ITS) sequence;
molecular identification;
adulterants;
B. falcatum
- From:
Chinese Journal of Experimental Traditional Medical Formulae
2020;26(14):182-189
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To establish a molecular identification method for Bupleurum chinense seeds based on ribosomal DNA internal transcribed spacer (ITS) sequence, ensuring the species authenticity of the cultivated seeds of B. chinense. Method:A total of 59 seeds samples of B. chinense and its main cultivated species, marketed B. chinense were collected. The effect of different sampling amounts and different water bath conditions on DNA extraction quality of the seeds was investigated, a DNA extraction method for seeds of Bupleurum was established. Their ITS sequences were obtained by polymerase chain reaction (PCR) and bidirectional sequencing. In addition, 34 ITS sequences of main cultivated Bupleurum species, such as B. chinense, B. scorzonerifolium, B. falcatum and B. smithii, were downloaded from GenBank to enrich identification database of B. chinense seeds. The neighbor-joining (NJ) dendrogram were constructed by MEGA-X 10.0.5 software to investigate the the species identification ability of ITS sequences for B. chinense seeds. And DNA barcoding identification of marketed B. chinense seeds was conducted based on BLAST method and NJ dendrogram method. Result:In total, 59 ITS sequences were obtained. ITS sequences of B. chinense could be divided into six haplotypes, including seven variable sites. The NJ dendrogram indicated that all the haplotypes of B. chinense could form independent branches, which could be distinguished from other cultivated species of Bupleurum in the collected samples, and possessed the ability to identify species of B. chinense seeds. Based on ITS sequence barcoding identification, 3 of the 19 marketed B. chinense seeds were B. falcatum with a counterfeit rate of 15.8%. Conclusion:DNA barcoding technology based on ITS sequence can accurately and reliably identify B. chinense seeds and its adulterants, providing reference for the standardization construction of Chinese medicinal materials seeds.
