Method for identifying Peru squids and highly processed products thereof through loop-mediated isothermal amplification technique

A loop-mediated isothermal, Peruvian squid technology, applied in the field of temperature amplification detection, can solve the problem of not using Peruvian squid, etc., and achieve the effects of simple operation, short detection time, and good specificity

Inactive Publication Date: 2016-08-17
ZHEJIANG GONGSHANG UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is currently no species identification m

Method used

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  • Method for identifying Peru squids and highly processed products thereof through loop-mediated isothermal amplification technique
  • Method for identifying Peru squids and highly processed products thereof through loop-mediated isothermal amplification technique
  • Method for identifying Peru squids and highly processed products thereof through loop-mediated isothermal amplification technique

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0079] Embodiment 1, adopt two kinds of LAMP detection methods to randomly buy frozen squid products (frozen squid rings, frozen squid fillets, etc.) or other aquatic product category identification on the market, including the following steps:

[0080] ① Extraction of genomic DNA from the sample to be tested; Axygen Genomic DNA Mini-Extraction Kit was used to extract DNA from the sample and stored at -20°C.

[0081] ②Design of specific identification primers for Peruvian squid;

[0082] Specific primers are specifically as described in Table 1 above.

[0083] ③LAMP amplification system setting;

[0084] Genomic DNA is used as an amplification template, and the above-mentioned specific primers are used for real-time fluorescent LAMP detection or LAMP staining detection.

[0085] details as follows:

[0086] Real-time fluorescence LAMP amplification system settings:

[0087] The reaction system is: 20 μL reaction system containing 10 μL of reaction buffer, 0.4 μL of upstrea...

Embodiment 2

[0104] Embodiment 2, adopt above-mentioned two kinds of LAMP detection methods to carry out kind identification to the canned squid commodity that buys randomly on the market:

[0105] Randomly purchase canned squid marked as canned squid or canned squid in its ingredients from the market, use TAKARA meat product DNA extraction reagent (Cat#9178) to extract DNA from the sample, and store it at -20°C. The species were identified by two LAMP detection methods. Real-time fluorescent LAMP method is used for detection. If the detected gene has no specific amplification curve or the detection time is ≥30min, the judgment result is negative, and it can be determined that the sample does not contain Peruvian squid; If the time is less than 30 minutes, it can be judged that the sample result is positive, that is, it is judged that the sample contains Peruvian squid; it is detected by LAMP staining method, and the reaction solution is mixed with SYBR After the I dyes are mixed, if the...

Embodiment 3

[0111] Embodiment 3, adopt above-mentioned two kinds of LAMP detection methods to carry out species identification to the deep-processing squid commodity that buys randomly on the market:

[0112] Randomly purchased from the market marked as squid shreds, squid slices and products containing squid in the product ingredients, TAKARA meat product DNA extraction reagent (Cat#9178) was used to extract sample DNA, and stored at -20 °C. Real-time fluorescent LAMP method is used for detection. If the detected gene has no specific amplification curve or the detection time is ≥30min, the judgment result is negative, and it can be determined that the sample does not contain Peruvian squid; If the time is less than 30 minutes, it can be judged that the sample result is positive, that is, it is judged that the sample contains Peruvian squid; it is detected by LAMP staining method, and the reaction solution is mixed with SYBR After the I dyes are mixed, if the reaction solution remains or...

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Abstract

The invention discloses specific primers which are adopted in an LAMP detection method for species identification of Peru liquids or highly processed products thereof and comprise the upstream outer primer (F3), the downstream outer primer (B3), the upstream inner primer (FIP) and the downstream inner primer (BIP). The invention further provides a method for identifying the Peru squids and the highly processed products thereof through a loop-mediated isothermal amplification technique. The method comprises the following steps of 1 extracting of genome DNA in a fish sample to be detected; 2 designing of the Peru squid LAMP amplification primers; 3 setting of a real-time fluorescent LAMP system and an LAMP staining method system; 4 judging according to detection results of the real-time fluorescent LAMP system and the LAMP staining method system.

Description

technical field [0001] The invention relates to two loop-mediated isothermal amplification detection methods suitable for Peruvian squid and its deep-processing products, namely, methods for identifying squid or its deep-processing products by using real-time fluorescent LAMP method and LAMP staining method. Background technique [0002] Peruvian squid (Dosidicus gigas), also known as squid, is a species of Cephalopoda, Teuthida, and Ommastrephidae. From 2003 to 2012, the catch of Peruvian squid increased from 400,000 tons to 950,000 tons. It is the largest species of cephalopod caught and has high commercial value. In the local Chinese market, besides Peruvian squid, the most common squid are Argentine squid (Illex argentinus), Japanese sea squid (Todarodes pacificus) and North Pacific squid (Ommastrephes bartramii). Squid meat is delicious and rich in a variety of essential ingredients for the human body. In addition to being directly sold fresh, the squid on the market w...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6888C12Q1/6844C12Q2531/119C12Q2561/113C12Q2563/107C12Q2545/113
Inventor 冯俊丽戴志远叶剑沈清金仁耀
Owner ZHEJIANG GONGSHANG UNIVERSITY
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