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Real-time PCR (Polymerase Chain Reaction) detection method of buffalo components in food and feed

A buffalo and feed technology, applied in the fields of molecular biology and nucleic acid detection, can solve the problem of specific detection of buffalo components without detection effect, etc.

Inactive Publication Date: 2014-03-05
PLANTS & ANIMALS & FOOD TESTING QUARANTINE TECH CENT SHANGHAI ENTRY EXIT INSPECTION & QUARANTINE BUREAU
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are no reagents and methods for the specific detection of buffalo components with good detection effect

Method used

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  • Real-time PCR (Polymerase Chain Reaction) detection method of buffalo components in food and feed
  • Real-time PCR (Polymerase Chain Reaction) detection method of buffalo components in food and feed
  • Real-time PCR (Polymerase Chain Reaction) detection method of buffalo components in food and feed

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0073] Embodiment 1, the detection result of eukaryote specific primer 18S rRNA primer

[0074] All the DNA solutions extracted in this experiment were amplified with eukaryotic 18S rRNA specific primers, and a specific amplification band of 137bp could appear in all DNA solutions. The results showed that all the extracted DNA solutions were suitable for PCR testing.

Embodiment 2

[0075] Embodiment 2, real-time fluorescent PCR specific detection of buffalo components

[0076] The DNA samples of 45 kinds of animal and plant materials were detected by using the detection primers and probes of buffalo components. Among them, the detection results of the milk samples of Mora buffalo, Nili-Rafi buffalo, the first-generation hybrid buffalo between Mora buffalo and Guangxi buffalo, the low-generation hybrid buffalo between Mora buffalo and Guangxi buffalo, and the high-generation miscellaneous buffalo between Mora buffalo and Guangxi buffalo appeared Obvious amplification curve, but no obvious amplification in DNA samples of other 39 species, such as figure 1 . A summary of the test results can be found in Table 1.

[0077] Table 1. Specific detection results of buffalo components

[0078]

[0079]

[0080] The above results show that the detection method of the present invention has species specificity.

Embodiment 3

[0081] Embodiment 3, the real-time fluorescent PCR detection sensitivity of buffalo component

[0082] The buffalo DNA solution was serially diluted 10 times with the DNA solution of ordinary milk (Dome fresh whole milk imported from Germany, purchased from a Shanghai supermarket). Real-time fluorescent PCR test was performed on 10ng / μL, 1ng / μL, 0.1ng / μL, 0.01ng / μL, 0.001ng / μL and 0.0001ng / μL buffalo milk sample DNA solutions using real-time fluorescent PCR detection primers and probes. The experiment was repeated 6 times.

[0083] In 6 tests, amplification curves appeared in 10ng / μL, 1ng / μL, 0.1ng / μL, 0.01ng / μL, 0.001ng / μL buffalo DNA solutions, but no amplification occurred in 0.0001ng / μL buffalo milk sample DNA. increasing curve, such as figure 2 .

[0084] Experiments show that at the level of DNA concentration, the detection sensitivity of the method of the present invention is 0.001ng / μL buffalo milk sample DNA.

[0085] Buffalo milk samples with different contents ...

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Abstract

The invention relates to a real-time PCR (Polymerase Chain Reaction) detection method of buffalo components in food and feed. The invention firstly discloses a primer for specific identification of buffalo components, and the primer can generate specific amplification of DNA (Deoxyribonucleic Acid) containing buffalo components, and can not generate specific amplification of DNA containing no buffalo components. The invention also provides a simplified PCR detection method, which can be well applied to the identification of the buffalo components, and has good reproducibility and sensitivity.

Description

technical field [0001] The invention relates to the technical fields of molecular biology and nucleic acid detection. Specifically, the present invention relates to a real-time fluorescent PCR detection method and kit for buffalo components in food or feed. Background technique [0002] As a kind of high-grade nutritious food, buffalo milk products have gradually become the "new favorite" of people's consumption, and are known as the "king of milk". Buffalo milk, buffalo cheese, buffalo milk health care products and baby food series are high value-added dairy products, whose product value and nutritional value are several times higher than ordinary milk products. As a commercially valuable ingredient, buffalo milk has higher sugar content, milk protein and casein content than yellow milk, and is rich in minerals and trace elements, with high nutritional value. The buffalo frankincense is thick, rich in flavor and full in taste , is a good nutrition for all ages. Buffalo m...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12N15/11
Inventor 张舒亚李富威王赢袁辰刚曾庆坤林波
Owner PLANTS & ANIMALS & FOOD TESTING QUARANTINE TECH CENT SHANGHAI ENTRY EXIT INSPECTION & QUARANTINE BUREAU