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Molecular identification kit of meat product made of beef and application of molecular identification kit

A technology for molecular identification and meat products, which is applied in the field of animal molecular biology, can solve the problems of poor sequence-level specificity and achieve low cost and strong specificity

Active Publication Date: 2014-09-24
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the specificity of the target sequence is not excellent at the sequence level. The detection results of the primers designed for common cattle show amplified products in both cattle and sheep, and non-specific bands amplify in other species.

Method used

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  • Molecular identification kit of meat product made of beef and application of molecular identification kit
  • Molecular identification kit of meat product made of beef and application of molecular identification kit
  • Molecular identification kit of meat product made of beef and application of molecular identification kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] 1. Sample preparation and storage

[0056] 1.1 Sampling

[0057] Collect 1 g of common beef product samples and store them at -20°C for future use.

[0058] 1.2 DNA template preparation

[0059] The DNA template is prepared using the commonly used phenol-chloroform crude extraction method (see: Sambrook J, Fritsch E F, Mani Artis T. Molecular Cloning Experiment Guide [M]. 2nd Edition. Jin Dongyan, Li Mengfeng Translated. Beijing : Science Publishing House, 1999.465-467) or other recognized DNA extraction methods with the same efficacy, these methods are all reported common methods.

[0060] 2. Primer Design

[0061] The DNA sequences of the primers in this example are shown in Table 1 and SEQ ID NO: 2 and 3 in the sequence table.

[0062] Table 1 PCR amplification primers designed by the present invention

[0063] Primer name

Primer sequence

TNFR-F

5′-CAGTGAGGACATCAGCCTAGAGT-3′

TNFR-R

5′-CTGCTCCTAGCCATCAGTGGA-3′

[0064] T...

Embodiment 2

[0084] The assembly of embodiment 2 kit

[0085] 1. The reagent composition in the kit is as described in Table 2:

[0086] Table 2PCR reaction system (ie reaction reagent)

[0087]

[0088] 2. One copy of the operation manual containing the standard PCR method as mentioned above.

[0089] 3. See Table 3 for kit composition, storage and stability testing.

[0090] Table 3 kit composition, storage conditions and stability test of the present invention

[0091]

[0092]

[0093] After testing, the assembled kit of the present invention will not affect its use effect if it is stored at room temperature for 12 months.

[0094] The application method of the test kit of the present invention is as follows:

[0095] 1. According to the following PCR reaction system, add the sample to a 200 μL EP tube (see Table 4):

[0096] Table 4PCR reaction system loading components and dosage

[0097]

[0098] 2. Load samples according to the above PCR amplification conditions; ...

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Abstract

The invention belongs to the technical field of meat product molecular detection and particularly relates to a molecular identification method and kit of a meat product made of beef as well as application of the molecular identification kit. The molecular identification method comprises the following steps: (1) primer designing: designing a specific primer according to the sequence of the TNFRSF10A gene of a bos taurus, wherein the sequence of the specific primer is shown in SEQ ID NO: 2 and SEQ ID NO: 3. (2) creating a PCR amplified reaction method, that is, amplifying the genome DNA of the bos taurus through the specific primer so as to obtain amplified specific fragments; (3) analyzing the amplified PCR products, that is, analyzing the amplified PCR products through agarose gel electrophoresis; according to gel-imaging, observing whether target fragments appear or not, and judging whether the target fragments contain the components of the genome DNA of the bos taurus or not. The molecular identification method has the characteristics of simplicity, convenience, quickness, high specificity and relatively-low cost, and is applicable to grassroots units.

Description

technical field [0001] The invention relates to the field of animal molecular biology, in particular to a molecular identification method of beef-derived meat products, a special kit and its application. The molecular identification method and kit of the present invention are related to PCR reaction. Background technique [0002] With the increasing popularity and coverage of meat products, in order to pursue profits, the market is often flooded with the phenomenon of "selling dog meat", and the safety of the source of meat products has become one of the focus issues that consumers care about. Identifying the source of meat products and establishing a convenient detection method are the keys to solving this problem. At present, foreign countries have published detection methods on animal origin in feed, which use tRNALys on mitochondria and ATPase ATPase subunits 8 and 6 subunit 8 and subunit 6 sequences as the target sequences for detection. However, the specificity of th...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68
Inventor 刘榜孙亚奇张庆德
Owner HUAZHONG AGRI UNIV
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