Dual fluorescent quantitative PCR method for species identification of salmons and highly processed products of salmons

A dual fluorescence and product variety technology, applied in the field of dual fluorescence quantitative PCR detection, can solve the problems of low cost, messy sequencing results, and inapplicability of mixed sample identification, and achieve the effect of shortening the detection time and reducing the cost of reagents

Inactive Publication Date: 2015-11-11
ZHEJIANG GONGSHANG UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This method is low in cost and easy to operate, but it also has many disadvantages: 1) Due to the existence of intraspecies variation and the degeneracy of different codons, different individuals of the same species may have different restriction fragment digestion patterns; 2) In addition to raw fish Outside of slices, deep salmon products (such as canned salmon) are usually processed at high temperature (>115°C) for a long time (>15min) during processing, an

Method used

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  • Dual fluorescent quantitative PCR method for species identification of salmons and highly processed products of salmons
  • Dual fluorescent quantitative PCR method for species identification of salmons and highly processed products of salmons
  • Dual fluorescent quantitative PCR method for species identification of salmons and highly processed products of salmons

Examples

Experimental program
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Effect test

Example Embodiment

[0090] Example 1. Using the above-mentioned dual fluorescence quantitative PCR detection method to identify the types of salmon sashimi randomly purchased on the market:

[0091] 1) Design of specific identification primers and probes for Salmonidae and 4 common salmon species;

[0092] Details are described in Table 1.

[0093] 2) Extraction of genomic DNA from salmon samples to be tested;

[0094] The sashimi marked as salmon was randomly purchased from the market, and the DNA of the sample was extracted using the Axygen Genomic DNA Mini Extraction Kit, and stored at -20°C.

[0095] 3) Fluorescence quantitative PCR amplification system settings:

[0096] The species was identified by dual fluorescence quantitative PCR.

[0097] The dual fluorescence quantitative PCR system is: AceQ is contained in a total volume of 20 μL TM qPCRProbeMasterMix 10μL, salmon specific upstream and downstream primers (5μM) 0.4μL each, salmon specific probe (10μM) 0.2μL, salmonidae 16SrDNA specific upstream a...

Example Embodiment

[0111] Example 2. The above-mentioned dual fluorescence quantitative PCR detection method is used to identify the species of salmon floss randomly purchased on the market:

[0112] Randomly purchased products labeled as salmon floss from the market, and extracted DNA from samples using Tiangen Deep Processed Food DNA Extraction Kit (DP326), and stored at -20°C. The species was identified by dual fluorescence quantitative PCR. If the Salmonidae 16SrDNA detection gene does not produce an S-type amplification curve and / or the Ct value is greater than 30, the judgement result is negative, it can be judged that the salmon floss sample does not contain salmonid fish components; if the Ct value is less than 30, it can If the result of the sample is determined to be positive, it is determined that the sample contains salmonid components. If none of the 4 common salmon CR detection genes produces an S curve and / or the Ct value is greater than 30, the judgment result is negative, it can b...

Example Embodiment

[0119] Example 3. The above-mentioned dual fluorescence quantitative PCR detection method is used to identify the types of canned salmon products randomly purchased on the market:

[0120] Randomly purchased products labeled as canned salmon fish from the market, using Tiangen Deep Processed Food DNA Extraction Kit (DP326) to extract the sample DNA, and store it at -20°C. The species was identified by dual fluorescence quantitative PCR. If the Salmonidae 16SrDNA detection gene does not produce an S-type amplification curve and / or the Ct value is greater than 30, the judgement result is negative, it can be judged that the canned fish sample does not contain Salmonidae fish components; if the Ct value is less than 30, it can If the result of the sample is determined to be positive, it is determined that the sample contains salmonid components. If none of the 4 common salmon CR detection genes produces an S curve and / or the Ct value is greater than 30, the judgment result is negati...

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Abstract

The invention provides a dual fluorescent quantitative PCR method for species identification of salmons and highly processed products of the salmons, and discloses a method for detecting the species of the salmons and the highly processed products of the salmons through dual fluorescent quantitative PCR. The method includes the following steps that firstly, specific identification primers and probes of salmonidae and four kinds of common salmons are designed, wherein the four kinds of common salmons include atlantic salmons, rainbow trouts, dog salmons and sockeye salmons; secondly, genome DNA in a salmon sample to be detected is extracted; thirdly, a dual fluorescent quantitative PCR amplification system is set; fourthly, judgment is made according to real-time fluorescent PCR detection results. The effective method is provided for quick detection of species identification of the salmons and the highly processed products of the salmons, and a technological guarantee is provided for aquatic product and food import and export.

Description

technical field [0001] The invention relates to a double fluorescent quantitative PCR detection method suitable for identification of salmon and its deep-processed products. Background technique [0002] Salmon is a general term for salmonid fish or salmon trout, and is known as "the supreme fish" and "treasure in water". Salmon is mainly produced in the border waters of the Arctic Ocean, the Atlantic Ocean, and the Pacific Ocean, and belongs to migratory fish in cold waters. Norway, which borders the Arctic Ocean, is the world's largest salmon farming and fishing country, accounting for half of the world's total production. Norwegian salmon is known as the "Emperor of the Ice Sea" by the international food industry for its best quality and best taste. At present, the domestically recognized economic species are roughly divided into 3 subfamilies, 7 genera, and about 36 species. As a commodity name, salmonids can be called salmon as long as the muscles are firm and elasti...

Claims

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Application Information

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IPC IPC(8): C12N15/11C12Q1/68
Inventor 冯俊丽戴志远吴志刚谢潇刘莎莎
Owner ZHEJIANG GONGSHANG UNIVERSITY
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