DNA Extraction and Molecular Identification in Angelicae Sinensis Radix Preparations

GOU Hui; WANG Yi-wei; ZHENG Xi; WANG Qin; ZHANG Chun

doi:10.13422/j.cnki.syfjx.2018010044

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DNA Extraction and Molecular Identification in Angelicae Sinensis Radix Preparations

更新时间：2024-01-04

- DNA Extraction and Molecular Identification in Angelicae Sinensis Radix Preparations
- Chinese Journal of Experimental Traditional Medical Formulae Vol. 24, Issue 1, Pages: 44-50(2018)
- 作者机构：
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- 基金信息：
- DOI：10.13422/j.cnki.syfjx.2018010044
  CLC： R284.1
- Published：2018
- 稿件说明：
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GOU Hui, WANG Yi-wei, ZHENG Xi, et al. DNA Extraction and Molecular Identification in Angelicae Sinensis Radix Preparations[J]. Chinese journal of experimental traditional medical formulae, 2018, 24(1): 44-50.
DOI：

GOU Hui, WANG Yi-wei, ZHENG Xi, et al. DNA Extraction and Molecular Identification in Angelicae Sinensis Radix Preparations[J]. Chinese journal of experimental traditional medical formulae, 2018, 24(1): 44-50. DOI： 10.13422/j.cnki.syfjx.2018010044.

摘要

目的：优化含当归中成药的DNA提取方法，并利用叶绿体trnL-F基因及当归特异分子标记对当归中成药进行鉴定。方法：采用传统十六烷基三甲基溴化铵（CTAB）法，十二烷基磺酸钠（SDS）法，磁珠法以及改良CTAB法对10种含有当归成分的中成药进行DNA提取，利用微量紫外分光光度法、琼脂糖凝胶电泳检测所得总DNA的完整性、纯度和浓度。对10份当归中成药的trnL-F序列进行扩增、测序，对序列进行比对分析并构建系统发育树。利用当归特异鉴别分子标记对10种当归中成药中的当归成分进行分子鉴别。结果：利用改良CTAB法获得了纯度较高，质量较好的DNA。10种中成药中当归的trnL-F序列与正品当归序列相似度为99.88%~100%，系统发育树显示10份中成药中的当归与正品当归聚为一类。利用当归特异性鉴别分子标记进行验证，10种中成药DNA样品均能得到明亮的扩增条带。结论：改良CTAB法可以有效提取当归中成药中的基因组DNA。利用trnL-F序列和当归特异性鉴别分子标记，成功对10份市售当归中成药进行了分子鉴别，为当归类中成药的分子鉴定奠定了基础。

Abstract

Objective:To optimize the DNA extraction method and molecular identification of Angelicae Sinensis Radix preparations by analyzing trnL-F sequences and using the Angelicae Sinensis Radix specific marker. Method:DNA was extracted by the traditional cetyltrimethylammonium bromide (CTAB) method

sodium dodecylsulphate (SDS) method

paramagnetic particle method and modified CTAB method from 10 kinds of Angelicae Sinensis Radix preparations. The integrity

purity

and concentration of the total genomic DNA yielded by the four methods were detected by ultraviolet spectrophotometry and agarose gel electrophoresis. The plastid trnL-F of 10 samples were amplified

sequenced and analyzed

and the phylogenetic trees were constructed. The DNA samples were also applied for molecular identification with Angelicae Sinensis Radix specific marker. Result:The modified CTAB method was found to yield relatively pure and high quality DNA from the 10 kinds of Angelicae Sinensis Radix preparations. The similarity of 10 samples and Angelicae Sinensis Radix ranged from 99.88% to 100%. Phylogenetic tree based on trnL-F sequences showed that 10 samples and Angelicae Sinensis Radix were clustered as one category. The clear and bright amplified band was obtained for all 10 samples in the verification by Angelicae Sinensis Radix specific molecular marker. Conclusion:The genomic DNA in Angelicae Sinensis Radix preparations could be successfully extracted by the modified CTAB method. The A. sinensis were successfully identified by the trnL-F sequences and the Angelicae Sinensis Radix specific molecular marker

with great importance in the molecular identification and quality control in Angelicae Sinensis Radix preparations.

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