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Real time fluorescent PCR detecting primer and probe for food sensibiligen shrimp component, kit and detecting method

A technology of sensitizing raw shrimp and real-time fluorescence, which is applied in biochemical equipment and methods, fluorescence/phosphorescence, material excitation analysis, etc., can solve the problems of pollution and low specificity, and achieve the effect of preventing pollution

Inactive Publication Date: 2012-08-22
曹际娟 +3
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  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

However, the specificity of conventional PCR is lower than that of real-time fluorescent PCR, and there are pollution problems

Method used

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  • Real time fluorescent PCR detecting primer and probe for food sensibiligen shrimp component, kit and detecting method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] 1. Design of primers and probes for real-time fluorescent PCR detection of food allergenic shrimp components

[0037] According to the results of homologous gene screening and comparison, housekeeping genes shrimp 16S rRNA gene I (SEQ ID NO.10), shrimp 16S rRNA gene II (SEQ ID NO.11) and arctic shrimp 16S rRNA gene ( SEQ ID NO.12) is the target gene for detection amplification. According to the nucleic acid sequences of the above-mentioned genes published on NCBI, and through comprehensive consideration of factors such as the consistency of the annealing temperature of the detection primers and probes and the similarity of GC content, the following specific primers and probes were designed:

[0038] ① The first pair of detection primers and probe sequences of shrimp 16S rRNA gene I:

[0039] Forward primer (SEQ ID NO.1):

[0040] 5′-GATTAAGTTACTTTAGGGATAACAGCG-3′,

[0041] Reverse primer (SEQ ID NO.2): 5'-TAAAGGTCGAACAGACCTTCTCACT-3',

[0042] Probe (SEQ ID NO.3): ...

Embodiment 2

[0098] Embodiment 2, specificity test

[0099] The detection kit and detection method containing specific primers and probes in Example 1 were used to detect shrimp allergen samples. Simultaneous detection of tiger head crab, sea crab, red crab, shrimp crawling, leek, rape, garlic, whitebait, flounder, yellow croaker, chives, red pepper, horseradish, carp, crucian carp, mandarin fish, cod, herring , wheat, salmon, purple stone house clam, Jiangyao shellfish, octopus and other non-shrimp allergen samples. To assess the specificity of the method.

[0100] Test result: no fluorescent signal is detected in the FAM channel of the negative control test result, indicating that the reaction result is normal, and the reaction system is pollution-free; the FAM channel of the positive control test result has a fluorescent signal detected, and the reaction result is normal; adopt the present invention to detect shrimp sensitization For the original samples, the fluorescence curve amplif...

Embodiment 3

[0101] Embodiment 3, shrimp allergen detection sensitivity test in raw material product

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Abstract

The present invention relates to real time fluorescent PCR detecting primer and probe for food sensibiligen shrimp component, a kit and a detecting method. The real time fluorescent PCR detecting primer and probe are respectively detecting primer and probe of shrimp 16 S / r RNS gene I of which the sequence is SEQ / ID / NO. 1 to 3, detecting primer and probe of shrimp 16 S / r RNS gene II of which the sequence is SEQ / ID / NO. 4 to 6, and detecting primer and probe of arctic shrimp 16 S / r RNS gene of which the sequence is SEQ / ID / NO. 7 to 9. On the basis of this, the present invention discloses the real time fluorescent PCR detecting kit for food sensibiligen shrimp component and the corresponding detecting method. The real time fluorescent PCR detecting primer and probe for food sensibiligen shrimp component have strong specificity; the real time fluorescent PCR detecting kit and method for food sensibiligen shrimp component is simple and easy; the result is exact; and real time fluorescent PCR detecting kit and method for food sensibiligen shrimp component have high specificity and flexibility.

Description

technical field [0001] The invention relates to a gene detection method for food allergen components. Especially the real-time fluorescent PCR detection method of food allergen raw shrimp components and the specific primers, probes and related kits involved therein. Background technique [0002] Studies have shown that food allergic reactions account for about 3.5% to 5% of people in developed countries. Even if these allergic people ingest foods containing trace allergens, they will also have very serious allergic reactions, even fatal. In addition, the tolerance of lactose and wheat protein also deserves the attention of allergic people. Allergic persons must avoid foods that contain relevant allergens or that exceed tolerated limits; therefore, any food containing a potentially sensitizing reaction must be properly labeled. The European Union's 2003 / 89 / EC directive on labeling requirements for food ingredients stipulates that any product containing allergens covered by ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12N15/11G01N21/64
Inventor 曹际娟郑秋月徐静徐君怡
Owner 曹际娟
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