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A Primer Set for Simultaneously Detecting Species Components of Blue Fox, Raccoon and Dog and Its Application

A primer set and species technology, applied in recombinant DNA technology, microbial determination/inspection, biochemical equipment and methods, etc., can solve the problems of less identification and detection, lack of effective detection and differentiation of canine components, etc., and achieve low cost. , good potential economic and social benefits, high specificity effect

Active Publication Date: 2016-11-02
INST OF SPECIAL ANIMAL & PLANT SCI OF CAAS
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in the research on the identification and detection of animal-derived components by molecular biological methods, there are few studies on the identification and detection of blue fox and raccoon-derived components, especially the lack of methods that can simultaneously and effectively detect and distinguish canine components

Method used

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  • A Primer Set for Simultaneously Detecting Species Components of Blue Fox, Raccoon and Dog and Its Application
  • A Primer Set for Simultaneously Detecting Species Components of Blue Fox, Raccoon and Dog and Its Application
  • A Primer Set for Simultaneously Detecting Species Components of Blue Fox, Raccoon and Dog and Its Application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Example 1 Extraction and Primer Design of Animal Blue Fox, Raccoon and Dog Mitochondrial Genomic DNA

[0025] Take 200 mg of muscle tissue from three kinds of animals respectively, and use the mitochondrial genomic DNA extraction kit, purchased from Beijing Baihao Biotechnology Co., Ltd. The extracted DNA was dissolved in TE buffer and stored at -20°C for later use.

[0026] 1. Sample Processing

[0027] a. Tissue homogenate: Weigh 100-200 mg of fresh tissue such as liver, brain, myocardium, etc., rinse with PBS or normal saline, wash the blood, blot dry with filter paper, cut into pieces with scissors and put them into a small-capacity glass homogenizer . Add 1.0 mL of ice-cold cell lysate, and grind the tissue up and down in an ice bath at 0°C for 20 times;

[0028] 2. Transfer the tissue or cell homogenate to a centrifuge tube, centrifuge at 1000×g for 5 minutes at 4°C;

[0029] 3. Take the supernatant, add it to a new centrifuge tube, and centrifuge again at 100...

Embodiment 2

[0041] Embodiment 2: PCR amplification and amplification product identification

[0042] The extracted blue fox, raccoon dog and canine mitochondrial genomic DNA were amplified by PCR;

[0043] The reaction system for PCR detection is 50 μL: 3 μL of extracted sample DNA template; 1.0 μL of each of the upstream primer and downstream primer; 25 μL of rtaq mix 2x enzyme, and 25 μL of pure water.

[0044] The PCR reaction program was as follows: pre-denaturation: 94°C, 3min; entry cycle: denaturation at 94°C for 60s, annealing at 58°C for 60s, extension at 72°C for 60s, 35 cycles; termination extension: 72°C, 7min; storage of the reaction product at 4°C.

[0045] PCR product electrophoresis detection and product sequence analysis: PCR products were electrophoresed in 1.0% agarose gel, and after 15 minutes, the gel imaging system was observed, and the three species of Canidae could be initially distinguished and identified by the size of the species-specific bands animal.

[0046...

Embodiment 3

[0047] Example 3 Simultaneous detection of blue fox, raccoon dog and dog species components in meat products

[0048] The market buys mutton, beef, pork, chicken, and dog meat; the experimental station collects muscle samples from fur animals such as blue foxes and raccoon dogs. The mitochondrial genome of each sample was extracted and detected by electrophoresis. And use a spectrometer and a nucleic acid analyzer to detect the concentration and purity. Samples that meet the requirements are frozen and stored for subsequent experiments.

[0049] 1. Single PCR detection

[0050] The obtained template DNA was individually added to any pair of primers in the three animals, and the result was that no specific band could be amplified for any primer pair in the mutton, beef, pork, and chicken templates. The primers of blue fox, raccoon dog and dog all showed specific bands for their own species, but the primers of blue fox and raccoon dog failed to obtain specific bands in dogs. ...

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Abstract

The invention discloses a primer group for simultaneously detecting species components of a blue fox, a racoon dog and a dog and application of the primer group and belongs to the technical field of detecting the specificity of components of animal origins. The primer group comprises a pair of blue fox species component detection primers, a pair of racoon dog species component detection primers and a pair of dog species component detection primers. The primer group is capable of simultaneously detecting the species components of the blue fox, the racoon dog and the dog. Meanwhile, the invention also discloses a method for simultaneously detecting the species components of the blue fox, the racoon dog and the dog by utilizing the primer group and a kit containing the primer group. The method has the characteristics of high specificity, high sensitivity, simpleness, reliability, rapidness and safety and is suitable for detecting and identifying the species components of the blue fox, the racoon dog and the dog in meat products, feeds and fur products.

Description

technical field [0001] The invention relates to a primer set for simultaneously detecting species components of blue fox, raccoon dog and dog and an application thereof, belonging to the technical field of specific detection of animal source components. Background technique [0002] The blue fox, also known as the Arctic fox, is a fur animal belonging to the family Canidae and the genus Arctic fox. It has been a special breeding animal in my country for decades. The performance of blue fox fur is excellent, and it is favored by the fur market. In recent years, the market demand for blue fox fur products is very large, and the number of breeding is also expanding. Raccoon dogs belong to the raccoon family of the family Canidae, and their fur is the main product. After the skins are harvested in winter, blue fox meat and raccoon meat are often used in animal feed and even food. At present, in my country’s animal husbandry industry, the relevant management regulations on insp...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6888C12Q2600/16
Inventor 孙伟丽李光玉钟伟王卓杨雅涵
Owner INST OF SPECIAL ANIMAL & PLANT SCI OF CAAS
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