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DNA barcode identification method of medical and edible material spina date seed and its fake product

A technology of jujube seed and barcode, which is applied in recombinant DNA technology, DNA/RNA fragment, and microbial determination/inspection.

Active Publication Date: 2017-12-15
河北省中药材质量检验检测研究中心有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, Zhang Junwu, Li Junqing, etc. use appearance traits, microscopic identification, thin layer identification, and ultraviolet spectrum identification to identify the jujube kernel and its counterfeit products. Liu Ping et al. RAPD analysis of Jen and its mixed and counterfeit products[J]. PLA Pharmaceutical Journal, 2006, 22(1): 67.), however, these identification methods all have certain defects: appearance traits and microscopic traits have a great influence on the expertise of technicians. The requirements for technical ability are relatively high, and more rely on the experience of technicians; thin-layer identification and ultraviolet spectroscopy are essentially based on the presence or absence or amount of a certain chemical component, and medicinal materials are caused by planting, harvesting or processing. Changes in the content of chemical components can easily lead to unstable results of thin-layer identification and ultraviolet spectroscopy; RAPD fragments have unknown genome sources (nuclei or organelles) and nucleotide homology of bands with the same gel electrophoretic mobility, etc. It is easy to cause interference. In addition, the RAPD reaction conditions are complicated and strict, and the stability of the amplified product is difficult to control.

Method used

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  • DNA barcode identification method of medical and edible material spina date seed and its fake product
  • DNA barcode identification method of medical and edible material spina date seed and its fake product
  • DNA barcode identification method of medical and edible material spina date seed and its fake product

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0069] Example 1 Intraspecific Variation Analysis of Suanzaoren Medicinal Material

[0070] After the comparison of 20 sequences from different sources of Zianzioren, the length of the psbA-trnH sequence was 321bp, and the GC content was 26.3%. There are 8 haplotypes at positions 232, 246, 258, 261, 268, 274, and 281. 5 samples of haplotype H1 and H2, same as 06AZ820151126363 and 06AZ820151126366 respectively; 4 samples of H3 haplotype, same as 06AZ820151126375; 2 samples of H4 haplotype, same as 06AZ820151126372; H5, H6, H7, H8 haplotype samples Both are 1, respectively 06AZ820151112501, EU075108, 06AZ820151126365 and 06AZ820151126367 (Table 2). The genetic distance was calculated based on the K2P model. The average K2P distance within the species of the psbA-trnH sequence of Suanzaoren medicinal material was 0.003, and the maximum K2P genetic distance within the species was 0.017.

[0071] Table 2 Intraspecific variation sites of psbA-trnH sequence in Suanzaoren

[0072] ...

Embodiment 2

[0074] Example 2 Interspecies Variation Analysis of Suanzaoren Medicinal Material and Its Counterfeit Products

[0075] After alignment of 18 sequences from samples from different sources in C. yunnanensis, the length of psbA-trnH sequence was 349bp, and the GC content was 25.4%. The minimum inter-species K2P distance of the psbA-trnH sequences of Suanzaoren and its adulterant, C. dianthus, was 0.310, and the average inter-species K2P distance was 0.321. The minimum interspecific distance between Suanzaoren and its counterfeit products was between Suanzaoren and Hovenia dulcis, which was 0.091, and the average K2P genetic distance was 0.093. The minimum interspecific K2P genetic distances of the psbA-trnH sequences of Suanzaoren and its counterfeit products were greater than the maximum intraspecific K2P genetic distances (Table 3).

[0076] Table 3 Intraspecific genetic distance of Jujube jujuba and its interspecific genetic distance with counterfeit varieties

[0077]

Embodiment 3

[0078] Example 3 Systematic cluster analysis of Jujube Seed Medicinal Materials and its counterfeit products

[0079] Based on the psbA-trnH sequence, the NJ phylogenetic clustering tree of jujube seed and its counterfeit products was constructed (see Figure 5 ). In the picture, jujube seed and its counterfeit products are clustered into different branches, which can be clearly distinguished from the counterfeit products. C. yunnanensis, Hovenia dulcis, Lentil and Bauhinia all clustered into one branch, showing good monophyletic.

[0080] In the present invention, a total of 5 species including Jujube Seed and its common counterfeit products, Lizaoren, Hovenia dulcis, Lentil and Bauhinia are selected for DNA barcode research. The intraspecific and interspecific genetic distance analysis of Suanzaoren and its counterfeit products showed that the maximum intraspecific genetic distance of 0.017 was much smaller than the minimum interspecific distance of 0.091 between Suanzaore...

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Abstract

The invention relates to a method for identifying authenticity of spina date seeds. The method comprises 1) extracting DNA of a sample to be tested, carrying out PCR amplification on the extracted DNA sequence and carrying out CodonCode Aligner splicing to obtain a psbA-trnH sequence, 2) carrying out comparison on K2P genetic distances and system clustering trees of the obtained psbA-trnH sequence and a psbA-trnH sequence of spina date seeds, and 3) If the test sample intraspecific maximum K2P genetic distance is less than the minimum interspecific K2P genetic distance from a fake product or the compared products are the same based on system clustering tree comparison, determining that the sample to be detected is true.

Description

technical field [0001] The invention belongs to the technical field of molecular identification, in particular to a DNA barcode identification method for wild jujube kernels and counterfeit products thereof Background technique [0002] Ziziphus jujube Mill.var.spinosa (Bunge) HuexH.F.Chou is the dry mature seed of Ziziphus jujube Mill.var.spinosa (Bunge) HuexH.F.Chou. It was first recorded in "Shen Nong's Materia Medica", listed as the top grade, and included in the previous editions of "The Pharmacopoeia of the People's Republic of China". It is mainly produced in northern my country. Efficacy, one of the traditional Chinese medicines commonly used in Chinese medicine clinics. Because of its health benefits such as helping sleep, lowering blood pressure, lowering blood fat, protecting the heart, and protecting the eyes, taking it for a long time can "safety the five internal organs, lighten the body and prolong life". Widely used in medicine and health food. However, wit...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/686C12Q1/6895C12Q2600/13C12Q2563/185C12Q2565/125
Inventor 张雅琴冯红马孝熙凃媛张兰兰
Owner 河北省中药材质量检验检测研究中心有限公司
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