Real-time fluorescence PCR (Polymerase Chain Reaction) detection method for source component of marten in food and feed

A component detection, real-time fluorescence technology, applied in biochemical equipment and methods, microbial measurement/inspection, etc., can solve the problems of ensuring the stability and specificity of the method, and achieve easy promotion, good stability and specificity, The effect of quick judgment

Inactive Publication Date: 2014-05-21
郑秋月 +3
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the detection method of mink-derived components in food and feed in my country is still blank, and there are many difficulties in ensuring the stability and specificity of the method, which is also the bottleneck restricting the establishment of the method

Method used

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  • Real-time fluorescence PCR (Polymerase Chain Reaction) detection method for source component of marten in food and feed
  • Real-time fluorescence PCR (Polymerase Chain Reaction) detection method for source component of marten in food and feed
  • Real-time fluorescence PCR (Polymerase Chain Reaction) detection method for source component of marten in food and feed

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] (1) Samples to be tested: Positive control, negative control, and blank control were set up during the testing process. DNA extracted from mink component samples was used as positive control, DNA extracted from samples known not to contain mink-derived components was used as negative control, sterilized water was used as blank control, and two parallel reaction systems were set up for each group.

[0054] (2) Detection method

[0055] (1) DNA extraction

[0056] The following DNA extraction methods are all routine operations of those skilled in the art, and all reagents and solutions are prepared by conventional means or obtained from commercial means.

[0057] ①Weigh 300mg of the prepared sample into a 2mL centrifuge tube, add 1.5mL CTAB buffer (55mmol / L CTAB, 1400mmol / L NaCl, 20mmol / L EDTA, 100mmol / L Tris, adjust the pH to 8.0 with 10% hydrochloric acid , 121°C, autoclave for 20min, spare.) and 10μL proteinase K (20mg / μL), shake and mix, 65°C for 30min, shake and mi...

Embodiment 2

[0089] Embodiment 2 specificity test

[0090] Use this standard method to test samples (see Table 2~3 for sample details). The sources of samples are Dalian Forest Zoo, Dalian Mink Breeding Farm, Dalian Fur Animal Breeding Base, and major supermarkets and markets in Dalian, including: Sable hair and meat, yellow-throated mink hair and blood, Japanese mink hair and blood, mink hair and meat, fox meat, cat meat and hair, weasel hair, horse meat, donkey meat, dog meat, mutton, beef, pork, chicken, Duck, goose, quail, tiger, wolf, leopard and other inbred or non-inbred DNA samples.

[0091] Partial results of specific detection of mink-derived components are as follows: Figure 4-5 And shown in Table 2~3.

[0092] Table 2 Results of detection specificity test of mink-derived components

[0093] serial number

sample name

test results

serial number

sample name

test results

1

mink fur

+

12

horsemeat

-

2

mink m...

Embodiment 3

[0100] Embodiment 3 detection sensitivity and addition test

[0101] Addition experiment was done with mutton as the substrate. After mink meat and mutton samples were taken respectively and ground in a meat grinder, the mink minced meat was added to the mutton minced meat and mixed evenly to form a 10% added sample. Add this ratio to the new minced mutton until the formation of mink meat 200mg / kg, 100mg / kg, 50mg / kg, 20mg / kg, 10mg / kg a total of 5 gradient addition samples. Mix well during addition to ensure even distribution of the sample throughout the lamb.

[0102] According to the result judgment standard described in embodiment 1, the amplification curve is judged; Add test result: as Image 6 The sensitivity test results of mink-derived ingredients addition experiments show that mutton is used as the base of processed products, mink meat is added respectively, and the added concentrations are 200mg / kg, 100mg / kg, 50mg / kg, 20mg / kg, 10mg / kg (corresponding to Image 6 1.2...

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Abstract

The invention discloses a primer and a probe for detecting a source component of marten as well as a use method and an application thereof. Aiming at cytochrome oxidase b gene of marten, the method designs the primer and probe for detecting the source component of marten; and components of marten in food and feed are quantitatively detected by a real time PCR (Polymerase Chain Reaction) method. The real-time fluorescence PCR quantitative detection method for components of marten in food and feed is of significance to improve the detection efficiency and agility of the source component of marten in food, lower the detection cost, detect related bogus products in market, and monitor safety of food and improve the inspection and quarantine technology. The method has a wide development prospect.

Description

technical field [0001] The invention belongs to the field of food quality control, in particular to a method for detecting mink-derived components in food and feed and its application. Background technique [0002] Mink is mainly produced in the Northeast region of China. There are many varieties and it is a precious fur animal. Mainly include sable, yellow-throated mink, Japanese mink and so on. In my country's fur farms, mink is the main breeding species. Mink belongs to the small and precious fur animals of Mammalia, Carnivora, Mustelidae and Mustelus in animal taxonomy. In the wild state, there are two kinds of American mink (M.vison) and European mink (M.lutreola). Now all countries in the world artificially raise the descendants of American and European mink, and our country mainly breeds the descendants of American mink. [0003] In recent years, many farms have raised mink artificially for their fur, and the remaining mink meat is often bought by unscrupulous trad...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68
Inventor 郑秋月徐君怡孙杰曹际娟
Owner 郑秋月
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