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SNP (single nucleotide polymorphism) molecular marker for identifying taxus cuspidata and application of SNP molecular marker

A molecular marker, yew technology, applied in the field of molecular biology, can solve the problems of subjective discrimination bias and morphological differences, and achieve simple operation, high specificity and high sensitivity, and objective results in 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

[0005] Morphological identification is a commonly used method in the identification of Northeast yew, 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 accurately identify only by morphological differences

Method used

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  • SNP (single nucleotide polymorphism) molecular marker for identifying taxus cuspidata and application of SNP molecular marker
  • SNP (single nucleotide polymorphism) molecular marker for identifying taxus cuspidata and application of SNP molecular marker
  • SNP (single nucleotide polymorphism) molecular marker for identifying taxus cuspidata and application of SNP molecular marker

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0062] Example 1 Obtaining of Taxus chinensis SNP molecular markers

[0063] 1. According to the t r n gene disclosed in the NCBI database, design corresponding primers for PCR amplification, using the upstream primer F1: 5'-GCAGAGACTCAACGGAAGAT-3' (SEQ ID NO.2), and the downstream primer R1: 5'-CCCTATTTATCATCCAAGCG-3' (SEQ ID NO.3), and the following PCR reaction system and PCR reaction program, with the identified southern yew (Taxus mairei SY Hu), Mandia yew (Taxus madia), European yew (Taxusbaccata), The genomic DNA of six taxus species from Tibet (Taxus wallichiana Zucc.), northeast yew (Taxus cuspidata S.etZ.) and Yunnan yew (Taxus yunnanensis Cheng et L.K.F) were used as templates for PCR amplification. The product was detected by 1.5% agarose gel electrophoresis and sequenced after being observed by the ultraviolet gel imaging system.

[0064]

[0065]

[0066] Among them, Vazyme AceTaq DNase (product number p401-d1 250U).

[0067] Extract plant genomic DNA acc...

Embodiment 2

[0071] Example 2 Application of Taxus chinensis SNP Molecular Marker in the Identification of Taxus chinensis by Fluorescent PCR Amplification

[0072] In this example, on the basis of obtaining the SNP site of Taxus chinensis, specific upstream primer F2, downstream primer R2 and probe P2 were designed for the gene sequence containing the SNP site using PrimerPremier5.0, and the primers and The probe was synthesized by Suzhou Jinweizhi Biotechnology Co., Ltd. The specific primers and probes are used to carry out fluorescent PCR amplification detection on Taxus chinensis, Taxus mandia, Taxus chinensis, Taxus tibetan, Taxus chinensis and Taxus yunnanensis. Specifically, the sequence of the upstream primer F2, the sequence of the downstream primer R2 and the sequence of the probe P2 are respectively:

[0073] Upstream primer F2: 5'-AGGTTGATCGTAAGATCAACTCATAT-3' (SEQ ID NO.4)

[0074] Downstream primer R2: 5'-AATTGTATGTATAATACAATTATAC-3' (SEQ ID NO.5)

[0075] Probe P2: 5'-TTA...

Embodiment 3

[0085] Example 3 Application of Northeast Taxus SNP Molecular Marker in Identification of Northeast Taxus by ARMS-PCR

[0086] In this example, on the basis of obtaining the SNP site of Taxus chinensis, specific upstream primer F3 and downstream primer R3 were respectively designed for the gene sequence containing the SNP site by using PrimerPremier5.0. Suzhou Jinweizhi Biotechnology Co., Ltd. Ltd Synthetics. The ARMS-PCR method is used to detect the samples of Taxus chinensis, Taxus Mandia, Taxus Europe, Taxus Tibet, Taxus Northeast China and Taxus Yunnan by using the specific primers. Specifically, the sequence of the upstream primer F3 and the sequence of the downstream primer R3 are respectively:

[0087] Upstream primer F3: 5'-AGGTTGATCGTAAGATCAACTCACAT-3' (SEQ ID NO.7)

[0088] Downstream primer R3: 5'-CAGAACCTGAGCTCAGGTCAACT-3' (SEQ ID NO.8)

[0089] Using the genomic DNA of 6 taxus species as templates, the 20 μL PCR reaction system includes:

[0090]

[0091] ...

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Abstract

The invention discloses an SNP (single nucleotide polymorphism) molecular marker for identifying taxus cuspidata. The SNP molecular marker is positioned at a 444th-site basic group of a sequence shownas SEQ ID NO.1. The taxus cuspidata is identified if the 444th-site basic group is T. The SNP molecular marker has the advantages that systems for quickly identifying the taxus cuspidata can be quickly established on the basis of the SNP molecular marker, accordingly, the purpose of quickly and accurately detecting the taxus cuspidata can be achieved, and the SNP molecular marker has important significance in port species inspection work and the like.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to a SNP molecular marker for Taxus chinensis and its application. Background technique [0002] Northeast yew (scientific name: Taxus cuspidata S.et Z.), also known as yew. It is a tree plant of Taxaceae (Taxaceae) and is a precious tree species left over from the Tertiary Period. Its material is excellent, straight texture, compact structure, high elasticity, high mechanical strength, luster, fragrance, corrosion resistance, not easy to crack and warp, does not contain turpentine, narrow yellowish white sapwood, purple reddish brown heartwood (yew is therefore named). This species is not only a timber plant, but also an important medicinal plant. Its stems, branches, leaves, and roots can be used as medicine, and its main components contain paclitaxel, taxine, and diterpenoids. It has anti-cancer function, and has the effect of inhibiting diabetes and treating heart disease. I...

Claims

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Application Information

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IPC IPC(8): C12Q1/6895C12N15/11
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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