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Reagent kit for bovine-derived component identification and detection of multi-species provenance components in products of bovine-derived components

A detection kit and kit technology, which is applied in the field of identification of bovine-derived components and detection kits for multi-species origin components in products, can solve the problems of high requirements, high cost, time-consuming and labor-intensive, etc., and achieve low cost, Wide applicability and strong specificity

Active Publication Date: 2016-01-27
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the fluorescence detection method has high requirements for equipment and is expensive; the enzyme digestion method is time-consuming and laborious; and the multiplex PCR is used to amplify the size of the fragment to identify the animal species, which severely limits the number of detectable species, and the primers often interfere with each other. Or form a dimer, affecting the reliability of the test results

Method used

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  • Reagent kit for bovine-derived component identification and detection of multi-species provenance components in products of bovine-derived components
  • Reagent kit for bovine-derived component identification and detection of multi-species provenance components in products of bovine-derived components
  • Reagent kit for bovine-derived component identification and detection of multi-species provenance components in products of bovine-derived components

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0070] Specific detection of bovine-derived components in the sample of Example 1

[0071] 1. Sample preparation and storage

[0072] 1.1 Sampling

[0073] Collect 1 g of beef samples and store them at -20°C for future use.

[0074] 1.2 DNA template preparation

[0075] The DNA template was prepared using the commonly used phenol-chloroform crude extraction method (Sambrook J, Fritsch EF, Maniartis T. Molecular Cloning Experiment Guide [M]. 2nd Edition. Jin Dongyan, Li Mengfeng. Beijing: Science Publishing Society, 1999.465-467) or recognized other extraction methods with the same efficacy, these methods are the commonly used methods reported.

[0076] 2. Primer Design

[0077] The DNA sequences of the primers in this embodiment are shown in Table 2 and SEQ ID NO: 2 and 3 in the sequence table.

[0078] Bovine PCR amplification primers designed by the present invention of table 2

[0079]

[0080] The size of the expected amplified fragment is 223bp, and its nucleotide ...

Embodiment 2

[0102] Embodiment 2 sensitivity test

[0103] Using 0.1ng / μL, 1ng / μL, 10ng / μL, and 100ng / μL of bovine DNA as templates, the nucleotide-specific fragment of SEQ ID NO.1 was amplified according to the conditions of Example 1. The result is as figure 2 As shown, there is amplification at a template concentration of 1ng / μL, indicating that the fragment amplified by the specific primer has good sensitivity.

Embodiment 3

[0104] Example 3 Detection of Primer Sets for Incorporating Non-Bovine Components in Bovine Products

[0105] The present invention is for the detection of animal-derived components. Through the PCR amplification of the corresponding primers of each species, it is judged whether it is or contains the DNA of a certain species, and then it is judged whether the animal-derived components of a certain species are mixed.

[0106] 1 Extraction of DNA from samples

[0107] The extraction and storage methods of the DNA of each species are the same as those described in Example 1 above.

[0108] 2 Primer design and screening

[0109] In order to find the specific detection sequence of each species, the present invention screens the specific sequence from the aspects of intra-species consistency and stability, inter-species specificity, copy number, etc., and compares it with the nucleic acid sequences of other species and different varieties of this species, The nucleotide sequences ...

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Abstract

The invention provides a reagent kit for bovine-derived component identification and detection of multi-species provenance components in products of bovine-derived components, and belongs to the technical field of animal provenance component molecular detection. The reagent kit comprises specific primers shown in the SEQ ID No.2 and the SEQ ID No.3, a reaction reagent and positive and blank control, and further comprises specific primers of pigs, horses, rabbits, foxes, dogs, minks, mice, chickens and ducks, and the nucleotide sequences of the specific primers are as shown from SEQ ID No.4 to SEQ ID No.21. By means of the primers of the reagent kit, sample DNA is amplified, a PCR amplification product is analyzed, whether a sample contains bovine-derived components or not can be judged and whether components of other animals such as pigs, horses, rabbits, foxes, dogs, minks, mice, chickens and ducks are doped in the sample or not can be judged. The reagent kit provides an efficient, precise and reliable means for detection of animal provenance components in meat, pelts and feed, and has the advantages of being easy and convenient to use, quick, comprehensive, high in specificity, low in cost and wide in adaptability.

Description

technical field [0001] The invention relates to the field of animal molecular biology, in particular to the identification of bovine-derived ingredients and a detection kit for multi-species origin ingredients in products thereof. Background technique [0002] In the consumption of animal food in our country, beef is second only to pork, and its nutritional value is high. There is an ancient saying that "beef nourishes qi, and its function is the same as that of astragalus". Beef is rich in protein, its amino acid composition is closer to the needs of the human body than pork, and its fat content is low, so it tastes delicious and enjoys the reputation of "the favorite of meat". Beef is rich in sarcosine, vitamins, carnitine, alanine and other nutrients, rich in iron, calcium, potassium, magnesium and other trace elements, and contains a fatty acid of conjugated linoleic acid, which can reduce fat deposition, There is a weight-loss effect, so beef products are favored by mo...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6888
Inventor 刘榜甄月然王文君付明张静刘建建吴清清向胜男张庆德
Owner HUAZHONG AGRI UNIV
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