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Dog endogenus DNA detection kit and identification method

A detection kit and identification method technology, applied in the field of dog-derived DNA detection kits and identification, can solve problems such as no dog meat, and achieve the effects of saving time, reliable detection results, and reducing costs

Inactive Publication Date: 2019-10-22
BEIHUA UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] At present, there is no legal identification method for dog meat in the national standards, and there are no relevant literature reports on the identification of dog meat, fox meat, and mink meat in China

Method used

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  • Dog endogenus DNA detection kit and identification method
  • Dog endogenus DNA detection kit and identification method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0058] 1. Pretreatment of test samples

[0059] The test samples were genuine dog meat (fresh), and the counterfeit products were fox meat (fresh) and mink meat (fresh), all of which were provided and identified by the Changchun Food and Drug Inspection Center. Take 0.05g fresh meat sample to be tested (fresh product), soak it in 1.5ml centrifuge tube with 0.9% Nacl aq, remove the connective tissue with a scalpel, mince the tissue, add 150ul Nacl aq to each tube, 10000r / min, 1min , discard the supernatant.

[0060] 2. Mitochondrial DNA Extraction

[0061] (1) Take the processed sample, add P1 (lysate, 0.5mol / L EDTA, 20mg / ml PK enzyme and 10mg / ml RNase) digestion solution 275μl, add P2 (1mol / L Tris-HCl, 0.5mol / L EDTA, NaCl, 10% SDS and 20mg / ml PK enzyme) lysate 250μl, mix carefully with a pipette tip, and place in a water bath at 56°C for 1h;

[0062] (2) After fully mixing, briefly centrifuge and place in a water bath at 70°C for 10 minutes;

[0063] (3) add to DNA purific...

Embodiment 2

[0075] Example 2 Identification of commercially available dog meat

[0076] 1. Materials

[0077] 3 samples of commercially available dog meat.

[0078] 2. Method

[0079] 2.1 Pretreatment of test samples Take 0.05g of fresh meat samples to be tested (fresh products), soak them in 0.9% Nacl aq in a 1.5ml centrifuge tube, remove the connective tissue with a scalpel and chop the tissue, add 150ul Nacl aq to each tube, 10000r / min, 1min, discard the supernatant.

[0080] 2.2 Mitochondrial DNA extraction

[0081] (1) Take the processed sample, add P1 (lysate, 0.5mol / L EDTA, 20mg / ml PK enzyme and 10mg / ml RNase) digestion solution 275μl, add P2 (1mol / L Tris-HCl, 0.5mol / L EDTA, NaCl, 10% SDS and 20mg / ml PK enzyme) lysate 250μl, mix carefully with a pipette tip, and place in a water bath at 56°C for 1h;

[0082] (2) After fully mixing, briefly centrifuge and place in a water bath at 70°C for 10 minutes;

[0083] (3) add to DNA purification column, centrifuge 3min at 10000rpm / min;...

Embodiment 3

[0094] Example 3 Identification of commercially available dog meat

[0095] The extraction steps and results of 6 batches of dog meat samples commercially available in Jilin City are the same as in Example 1.

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Abstract

The invention relates to the technical field of meat detection and discloses a dog endogenus DNA detection kit. The dog endogenus DNA detection kit is characterized by being composed of a sample pretreatment solution, a mitochondria DNA extraction system and a PCR system. The total system volume of the PCR system is 20 microliters, and the PCR system contains 10 microliters of Taq Plus PCR MasterMix, 2 microliters of a 10 mM dog meat primer, 1 microliter of to-be-detected sample DNA and 7 microliters of sterile double distilled water. An identification method for dog endogenus DNA comprises the steps of designing a dog meat specific oligonucleotides primer, determining a reaction procedure, judging a result and the like. According to a judgement standard, in a PCR amplification result, ifa 481bp strip exists, the salable product dog meat is determined, if multiple strips exist or the strip does not exist, the counterfeit product dog meat is determined, if a 393bp strip exists, the counterfeit product fox meat is determined, if a 289bp strip exists, the counterfeit product marten meat is determined, and reference is shown in a graph 1 and a graph 2. According to the identificationmethod, the true and false dog meat can be accurately identified.

Description

technical field [0001] The present invention relates to the use of molecular biology techniques to identify dog-derived and easily mixed species (fox, mink)-derived DNA and an identification method. Background technique [0002] Dog meat is the meat of canine dogs, also known as "fragrant meat", "ground sheep" and "white dog". Dog meat contains protein, fat, vitamin A, vitamin B1, vitamin B2, vitamin B3, vitamin C, calcium, iron, phosphorus, potassium, sodium, and Ash. The efficacy of dog meat nourishing qi and blood was first recorded in the "Compendium of Materia Medica" "passing the two meridians and instantly warming the stomach and dispelling cold, taking it to overflow the qi and blood, and make the pulse boil". [0003] In recent years, due to the serious reduction of dog meat resources, the price has been on an upward trend, resulting in the emergence of a large number of counterfeit and confused products, seriously disrupting the normal market order, especially fox...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6888C12Q1/686C12Q1/6806
CPCC12Q1/6888C12Q1/686C12Q1/6806C12Q2565/125
Inventor 孙丽媛王天添陈思秀刘玟妍关奕泽艾金霞李明成赵云冬王艳双高丽君王雪松
Owner BEIHUA UNIV