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SNP molecular marker for identifying European red bean fir or Taxus media red bean fir

A technology of Taxus mandia and molecular markers, which is applied in the direction of recombinant DNA technology, microbial measurement/inspection, DNA/RNA fragments, etc., can solve the problems of subjective discrimination bias and morphological differences, and achieve simple operation and high specificity and high sensitivity, the objective effect of the identification process

Pending Publication Date: 2019-03-19
HANGZHOU KMB BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Morphological identification is a commonly used method in the identification of Taxus chinensis or Taxus mandia, but the shape characteristics are easily affected by habitat, climate, physiological conditions, etc., which often lead to deviations in subjective identification, and it is difficult to identify accurately only by morphological differences

Method used

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  • SNP molecular marker for identifying European red bean fir or Taxus media red bean fir
  • SNP molecular marker for identifying European red bean fir or Taxus media red bean fir
  • SNP molecular marker for identifying European red bean fir or Taxus media red bean fir

Examples

Experimental program
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Effect test

Embodiment 1

[0060] Example 1 Obtaining of European or Taxus mandia SNP molecular markers

[0061] 1. According to the ITS gene disclosed in the NCBI database, design corresponding primers for PCR amplification, using upstream primer F1: 5'-GCGGTAGGATCATTGTCG-3' (SEQ ID NO.2), downstream primer R1: 5'-TCCTCCGCTTATTGATATGC-3' (SEQ ID NO.3), and the following PCR reaction system and PCR reaction program, with the identified southern yew (Taxusmairei SY Hu), Mandiya yew (Taxus madia), European yew (Taxus baccata), The genomic DNA of six species of Taxus genus, Taxus wallichiana Zucc., Taxus cuspidata S.et Z., and Taxus yunnanensis Cheng et L.K.F, were used as templates for PCR amplification. The amplified products were detected by 1.5% agarose gel electrophoresis and sequenced after being observed by the ultraviolet gel imaging system.

[0062]

[0063] Among them, Vazyme AceTaq DNase (product number p401-d1 250U).

[0064] Extract plant genomic DNA according to traditional methods, or u...

Embodiment 2

[0067] Example 2 Application of European or Mandia Taxus SNP Molecular Markers in Fluorescent PCR Amplification Method Identification of Europe or Mandia Taxus

[0068] In this example, on the basis of obtaining the SNP sites of Taxus chinensis or Mandia, specific upstream primers F2, downstream primers R2 and probes were designed using PrimerPremier5.0 for the gene sequence containing the SNP sites P2, primers and probes were synthesized by Suzhou Jinweizhi Biotechnology Co., Ltd. The specific primers and probes were used to carry out fluorescent PCR amplification detection on Taxus chinensis, Taxus Mandia, Taxus Europe, Taxus Tibet, Taxus Northeast China and Taxus Yunnan. Specifically, the upstream primer F2, downstream primer R2 and probe P2 are respectively:

[0069] Upstream primer F2: 5'-CCGGTCTGCAAAGCAGCT-3' (SEQ ID NO.4)

[0070] Downstream primer R2: 5'-GGAACCAGTGGACGGTCG-3' (SEQ ID NO.5)

[0071] Probe sequence P2: 5'-GATTATTTCGCGCCTTTGGACG-3' (SEQ ID NO.6)

[00...

Embodiment 3

[0081] Example 3 Application of European or Mandia Taxus SNP Molecular Markers in Arms-PCR Identification of Taxus Tibet

[0082] In this example, on the basis of obtaining the SNP sites of Taxus chinensis or Mandia, specific upstream primers F3 and downstream primers R3 were respectively designed for the gene sequence containing the SNP sites using PrimerPremier5.0. The primers consist of Synthesized by Suzhou Jinweizhi Biotechnology Co., Ltd. Utilize described specific primer and probe to carry out fluorescent PCR amplification detection to Southern Taxus, Mandia Taxus, European Taxus, Tibet Taxus, Northeast Taxus and Yunnan Taxus samples. Specifically, the sequence of the upstream primer F3 and the sequence of the downstream primer R3 are respectively:

[0083] Upstream primer F3: 5'-TAATGCCGTACAGATTTATTTCGTGC-3' (SEQ ID NO.7)

[0084] Downstream primer R3: 5'-GAGAGCCGAGATATCCGTTGC-3' (SEQ ID NO.8)

[0085] Using the genomic DNA of 6 taxus species as templates, the 20 μL...

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Abstract

The invention discloses an SNP molecular marker for identifying an European red bean fir or a Taxus media red bean fir, wherein the SNP molecular marker is positioned at a 450th basic group of the sequence shown in SEQ ID NO. 1, and if the 450th basic group is C, the SNP molecular marker is identified as the European red bean fir or the Taxus media red bean fir. The invention establishes a systemfor rapidly identifying the European red bean fir or the Taxus media red bean fir based on the SNP molecular marker, and thus the purpose of rapidly and accurately detecting the European red bean firor the Taxus media red bean fir is achieved, so that the SNP molecular marker is of great significance for species inspection at ports and the like.

Description

technical field [0001] The present invention relates to the field of molecular biology detection of wood tree species identification, in particular, relates to the SNP molecular marker of European or Mandia yew, and uses the SNP site to identify the above two types of tree species from other yew varieties Applications. Background technique [0002] Yew, also known as yew, belongs to the national first-class protected plants, and is a rare and endangered species in the world left over from the Quaternary Glacier. Taxus has both tall trees and low shrubs. There are about dozens of species in the world, and Taxus Mandia is one of them. Taxus Mandia is one of the yew species. Taxus mairei (scientific name: Taxus mairei) is a relic plant of the Quaternary Ice Age. It is a rare and endangered species in the world. There are eleven species in the world. At present, there are four species and one species in my country. Varieties, namely Taxus Yunnan, Taxus Tibet, Taxus Northeast Ch...

Claims

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Application Information

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IPC IPC(8): C12Q1/6895C12Q1/686C12N15/11
CPCC12Q1/686C12Q1/6895C12Q2600/156C12Q2561/113C12Q2563/107C12Q2545/114C12Q2535/137
Inventor 林源吉丁佳女郑宜文
Owner HANGZHOU KMB BIOTECH
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