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Kit for detecting swill-cooked dirty oil and detection method of kit

A technology of waste oil and kits, applied in the direction of biochemical equipment and methods, microbial determination/inspection, etc.

Active Publication Date: 2015-02-04
NORTHWEST UNIVERSITY FOR NATIONALITIES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there is no kit for detecting the characteristic DNA sequence and primer set of animal mitochondria by using the LAMP method to construct waste oil.

Method used

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  • Kit for detecting swill-cooked dirty oil and detection method of kit
  • Kit for detecting swill-cooked dirty oil and detection method of kit
  • Kit for detecting swill-cooked dirty oil and detection method of kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0080] Example 1 A test kit for waste oil detection, comprising

[0081] —Detection primer solution: composed of outer primer I with a concentration of 4~6 μmol / L, outer primer II with a concentration of 4~6 μmol / L, inner primer I with a concentration of 32~48 μmol / L and a concentration of 32~48 μmol / L / L internal primer II prepared;

[0082] - Bst DNA polymerase with strand displacement activity: the concentration is 7~9U / μL;

[0083] —10× reaction buffer: Tris-HCl of 200 mmol / L and pH=8.8, 100 mmol / L KCl, 100 mmol / L (NH 4 ) 2 SO 4 , 40~100 mmol / L MgSO 4 , mixed with 6~14 mol / L betaine;

[0084] —dNTPs solution: It is formed by mixing equal volumes of four deoxyribonucleotide solutions of dATP, dCTP, dGTP and dTTP with a concentration of 10 mmol / L;

[0085] —Positive DNA control sample: Escherichia coli plasmid DNA containing highly conserved DNA sequences in vertebrate mitochondria, or total DNA samples of pigs, cattle, sheep, chickens, ducks and fish commonly foun...

Embodiment 2

[0093] Example 2 A detection method of a kit for waste oil detection, comprising the following steps:

[0094] ⑴Extract the DNA of the sample to be tested:

[0095] Add 400μL TE buffer solution to 2mL centrifuge tube Ⅰ, then add 1mL edible oil, vortex and shake for 3~8min, centrifuge at 13000r / min at room temperature for 5min, remove the upper layer of oil, leave a layer of water phase, add again to centrifuge tube Ⅰ Edible oil 1mL, mix upside down, centrifuge, repeat 5-20 times in total, the obtained water phase is the DNA of the sample to be tested;

[0096] ⑵ Prepare the LAMP detection reaction system for the sample to be tested:

[0097] Add 2~5 μL of DNA to be tested, 1.5 μL of detection primer solution, 1 μL of Bst DNA polymerase, 2.5 μL of 10× reaction buffer, and 3 μL of dNTPs solution into 200 μL PCR reaction tube Ⅰ, and replenish with sterilized deionized water. Dilute to 25 μL;

[0098] (3) Prepare the LAMP detection reaction system for the control sample:

...

Embodiment 3

[0112] Example 3 A test kit for waste oil detection, comprising

[0113] —Detection primer solution: composed of outer primer I with a concentration of 4~6 μmol / L, outer primer II with a concentration of 4~6 μmol / L, inner primer I with a concentration of 32~48 μmol / L and a concentration of 32~48 μmol / L / L internal primer II prepared;

[0114] - Bst DNA polymerase with strand displacement activity: the concentration is 7~9U / μL;

[0115] —10×reaction buffer: 200 mmol / L Tris-HCl with pH=8.8, 100 mmol / L KCl, 100 mmol / L (NH 4 ) 2 SO 4 , 40~100 mmol / L MgSO 4 , 6~14 mol / L betaine;

[0116] —dNTPs solution: It is formed by mixing equal volumes of four deoxyribonucleotide solutions of dATP, dCTP, dGTP and dTTP with a concentration of 10 mmol / L;

[0117] —Positive DNA control sample: Escherichia coli plasmid DNA containing highly conserved DNA sequences in vertebrate mitochondria, or total DNA samples of pigs, cattle, sheep, chickens, ducks and fish commonly found in food; ...

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Abstract

The invention relates to a kit for detecting swill-cooked dirty oil. The kit comprises a detection primer solution prepared from an outer primer I, an outer primer II, an inner primer I and an inner primer II, a BstDNA polymerase with chain displacement activity, a 10*reaction buffer, a dNTPs solution, a positive DNA control sample and a negative DNA control sample. Meanwhile, the invention also discloses a detection method of the kit. The kit disclosed by the invention has the characteristics of high specificity, high speed, high efficiency and low detection cost, is simple and convenient to operate and is suitable for rapid field detection.

Description

technical field [0001] The invention relates to a detection kit, in particular to a detection kit for waste oil detection and a detection method thereof. Background technique [0002] Waste oil is a kind of non-edible oil with extremely poor sanitation, peroxide value, acid value, moisture, carbonyl value, malondialdehyde, aflatoxin, etc. seriously exceeding the standard. The main ways of production are as follows: one is from some greasy floating substances in the sewer, and some greasy substances in waste kitchen oil, which are extracted through simple processing; the other is extracted from tissues such as animal viscera Grease; the third is the fat that has been repeatedly fried. Long-term consumption of various toxic substances such as aflatoxin, benzo(a)pyrene, and trans fatty acid in waste oil is very harmful to human body and seriously threatens the health of human beings. In recent years, domestic reports about waste oil and the food safety problems caused by wast...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
CPCC12Q1/6844C12Q1/6876C12Q2531/119
Inventor 徐红伟臧荣鑫
Owner NORTHWEST UNIVERSITY FOR NATIONALITIES
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