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基于ITS2条形码鉴定野菊花及其混淆品

[Identification of Chrysanthemum indicum and its adulterants based on ITS2 barcode].

作者信息

Chen Fu-Rong, Wang Tao, Guo Qiao-Sheng, Zhu Zai-Biao, Zou Qing-Jun, Gui Shu-Qi, Zhao Shu-Yi

机构信息

Institute of Chinese Medicinal Materials,Nanjing Agricultural University Nanjing 210095,China.

出版信息

Zhongguo Zhong Yao Za Zhi. 2019 Feb;44(4):654-659. doi: 10.19540/j.cnki.cjcmm.20181204.010.

Abstract

DNA barcode technology was used to establish a rapid identification method of Chrysanthemum indicum based on ITS2 sequences. The total DNA was extracted from 22 collected samples,and the ITS2 sequence was amplified by PCR and sequenced,and the information of ITS2 sequence was obtained. Another 14 items of the same family or the same genus were downloaded from Gen Bank.We aligned all 36 sequences,calculated the intraspecific and interspecific distances,and constructed Neighbor Joining( NJ) phylogenetic tree,using MEGA 7. 0. The difference of the secondary structure between the ITS2 sequences was compared. The results showed that the genetic distance of Ch. indicum and Ch. morifolium was overlapped,but the maximum intraspecific distance was far less than the minimum interspecific distance between and among Ch. indicum and other species,with an obvious barcoding gap. The NJ tree showed that Ch. indicum and Ch. morifolium shared a clade,and most of Ch. morifolium with some Ch. indicum were shared a subclade,while Inula lineariifolia,Sinosenecio oldhamianus and Senecio scandens belonged to one clade separately. ITS2 secondary structures for I. lineariifolia,S. oldhamianus and S. scandens were significantly different enough to identify completely but Ch. indicum and Ch. morifolium shared two secondary structures of A and B. It was proved that Ch. indicum was one of the evolutionary sources of Ch.morifolium. Therefore ITS2 sequence as DNA barcode can identify Ch. indicum and its adulterants accurately and quickly. The study provides an important basis for Ch. indicum for the identification of germplasm resources and the safety of clinical medication.

摘要

采用DNA条形码技术,基于ITS2序列建立了野菊花快速鉴定方法。从采集的22份样品中提取总DNA,通过PCR扩增ITS2序列并测序,获得ITS2序列信息。从GenBank下载同科或同属的另外14条序列。利用MEGA 7.0对36条序列进行比对,计算种内和种间距离,构建邻接(NJ)系统发育树。比较ITS2序列二级结构的差异。结果表明,野菊花与菊花的遗传距离有重叠,但野菊花种内最大距离远小于其与其他物种间的最小种间距离,存在明显的条形码间隙。NJ树显示,野菊花与菊花共享一个分支,多数菊花与部分野菊花共享一个亚分支,而线叶旋覆花、少花紫菊和千里光分别属于一个分支。线叶旋覆花、少花紫菊和千里光的ITS2二级结构差异显著,足以完全鉴别,而野菊花与菊花共享A和B两种二级结构。证明野菊花是菊花的进化来源之一。因此,ITS2序列作为DNA条形码可准确快速地鉴别野菊花及其混淆品。该研究为野菊花种质资源鉴定及临床用药安全提供了重要依据。

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