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Kit for detection of animal origin components and detection method

A kit and animal-sourced technology, applied in biochemical equipment and methods, microbiological determination/inspection, etc., can solve the problems of high detection cost and achieve the effects of improved detection sensitivity, high degree of visualization, and strong specificity

Inactive Publication Date: 2017-11-28
SHANDONG INST FOR FOOD & DRUG CONTROL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Aiming at the current problem of high cost of detecting multiple animal-derived components at the same time, the present invention provides a kit for detecting multiple animal-derived components

Method used

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  • Kit for detection of animal origin components and detection method
  • Kit for detection of animal origin components and detection method
  • Kit for detection of animal origin components and detection method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1 Preparation of kit

[0037] 1.1 Preparation of membrane chip

[0038] The five probes of pig, sheep, yak, donkey, and mouse as shown in Seq No. 1-8 as well as positive probe, negative probe and internal reference probe are prepared into a solution with a concentration of 25 μmol / L with ultrapure water , Using a micro-quantitative spray-point membrane chip spotting instrument to distribute each nucleotide probe in a specific location area on the negatively charged nylon membrane chip. The layout of the probes coated on the chip surface is as follows figure 1 As shown, PC is a positive probe and NC is a negative probe.

[0039] 1.2 Reagent preparation

[0040] Prepare 10×PCR buffer (P1), containing 15 mmol / L MgCl 2 , 100 mmol / L Tris-HCl, 500 mmol / L KCl, and its pH is 8.3.

[0041] Prepare the multiple PCR primer master mix (P2) with the amplification primer pairs of pig, sheep, yak, donkey, and mouse as shown in Seq No. 10-21 and the internal reference amplification prim...

Embodiment 2

[0049] Example 2 Determination of the specificity and sensitivity of the kit for detecting 5 kinds of animal-derived components

[0050] 2.1 Extraction and purification of DNA in the experiment

[0051] After pre-processing the samples containing pig, sheep, donkey, yak, and mouse components, extract and purify DNA, and use a nucleic acid protein analyzer to detect its concentration and quality OD 260 / OD 280 When the value is between 1.7-1.9, it is used for multiplex PCR amplification. The DNA extracted from the corresponding animal-derived species was used as a positive control, the DNA of a species known to be free of the animal-derived species was used as a negative control, and sterilized water was used as a blank control.

[0052] 2.2 Multiplex PCR

[0053] Use the DNA obtained in 2.1 as a template to perform multiplex PCR. The system is shown in Table 1, and the reaction parameters are as follows to obtain the amplified product:

[0054] Firstly, it is denatured at 95°C for 10 m...

Embodiment 3

[0073] Example 3 Test kit for detecting ingredients in meat products

[0074] Collect 173 market samples from sources such as farmers’ markets, supermarkets, and the Internet. The sample types include fresh meat, meat skewers, meatballs, pre-made meat, jerky, ham sausage, etc. The methods described in 2.1-2.4 The samples were tested, and the results are shown in Table 3.

[0075] Table 3 Test results of animal-derived components of samples

[0076]

[0077] It was found that the test results of fresh meat samples had a relatively high compliance rate with their labels, about 85%, and there were frauds in yak meat, donkey meat, and mutton; 10 meat skewers with labels only labeled with sheep ingredients were all detected. However, 2 samples were detected with pig ingredients, and the test results were 80% in compliance with their labels; 17 meatball samples were labeled with animal-derived ingredients such as chicken, pig, sheep, and duck. Porcine-derived ingredients were found, of w...

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Abstract

The invention provides a kit for detection of animal origin components. The kit includes a membrane chip, the membrane chip is composed of a support membrane and probes fixed on the support membrane. The probes are specific probes for detection of pigs, sheep, yaks, donkeys and rats, the sequences of the probes are shown as Seq No.1-5, and the 5' end of each probe is labeled with amino. The kit also includes an internal reference probe for detection of internal reference gene, a positive probe and a negative probe, the sequences of the probes are shown as Seq No.6-8, and the 5' end of each probe is labeled with amino. The kit also includes positive oligonucleotide single stranded DNA with a biotin labeled 5' end, and the sequence is shown as Seq No.9. The kit provided by the invention can detect whether different varieties of meat products contain pigs, sheep, donkey, yak and rat 5 animal origin components substantially at a time, and the obtained detection result has the characteristics of strong specificity, high sensitivity, low false positive rate, high visual degree and low cost, etc.

Description

Technical field [0001] The invention belongs to the technical field of detection supplies, and in particular relates to a kit and a detection method for detecting multiple animal-derived components. Background technique [0002] In recent years, my country's food industry has entered a golden period of rapid development. Poultry and livestock meat products and their products have enriched the people's dining table, but meat products have frequently been adulterated. Incidents such as fox meat and duck meat pretending to be lamb, beef burger patties mixed with horse meat, etc., have become a hot and difficult issue in the field of food safety supervision. With the development of the modern meat product industry, in order to weigh the taste of meat products and the cost-effectiveness of production and processing, more and more meat products on the market are processed from various types of animal-derived meat, and the same sample often needs to be carried out at the same time. Det...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCC12Q1/6837C12Q1/686C12Q1/6888C12Q2537/143
Inventor 霍胜楠任易婕孟静郑世超伊廷存
Owner SHANDONG INST FOR FOOD & DRUG CONTROL
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