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Preparation and test method of rapid test kit for Roundup Ready transgenic Soybean and processed products thereof

A technology of genetically modified soybeans and detection kits, applied in biochemical equipment and methods, material analysis through observation of the impact on chemical indicators, microbial determination/inspection, etc., can solve problems such as expensive reagents and instruments

Inactive Publication Date: 2009-10-07
INSPECTION & QUARANTINE TECH CENT SHANDONG ENTRY EXIT INSPECTION & QUARANTINE BUREAU
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Currently, among DNA-based detection methods, polymerase chain reaction (PCR) is mainly used, which is sensitive and specific, but requires expensive reagents and instruments

Method used

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  • Preparation and test method of rapid test kit for Roundup Ready transgenic Soybean and processed products thereof
  • Preparation and test method of rapid test kit for Roundup Ready transgenic Soybean and processed products thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036]The loop-mediated isothermal amplification detection kit for glyphosate-resistant transgenic soybeans was prepared according to the following formula:

[0037] (1) Ring-mediated isothermal amplification reaction solution A

[0038] 10×Bst buffer reaction buffer 2.5 μL, 25 mmol / L dNTP 1.3 μL, 100 mmol / L magnesium sulfate 3 μL, 2mol / L betaine 5 μL, 10 μmol / L upstream internal primer A 3 μL, 10 μmol / L downstream internal primer A 3 μL, 10 μmol / L L upstream outer primer A 0.5 μL, 10 μmol / L downstream outer primer A 0.5 μL, ddH 2 O (sterilized double distilled water) 3.2 μL;

[0039] The sequence of the upstream internal primer A is: 5'-AGGTGGCACCTACAAATGCCATCGGCAGAGGCATCTTGAAC-3'; the sequence of the downstream internal primer A is: 5'-TAGCTGGGCAATGGAATCCGAGGTCTCCAGAAGACCAAAGGGC-3'; the sequence of the upstream external primer A is: 5'-TCCATCTTTTGGGACCACTGT-3'; The sequence of the outer primer A is: 5'-CGACACTCTCGTCTACTCCA-3';

[0040] The volume ratio of the upstream int...

Embodiment 2

[0066] The loop-mediated isothermal amplification detection kit for glyphosate-resistant transgenic soybeans was prepared according to the following formula:

[0067] (1) Ring-mediated isothermal amplification reaction solution A

[0068] 10×Bst buffer reaction buffer 2.5 μL, 25 mmol / L dNTP 1.3 μL, 100 mmol / L magnesium sulfate 3 μL, 2mol / L betaine 5 μL, 10 μmol / L upstream internal primer A 3 μL, 10 μmol / L downstream internal primer A 3 μL, 10 μmol / L L upstream outer primer A 0.5 μL, 10 μmol / L downstream outer primer A 0.5 μL, ddH 2 O (sterilized double distilled water) 3.2 μL;

[0069] The sequence of the upstream internal primer A is: 5'-AGGTGGCACCTACAAATGCCATCGGCAGAGGCATCTTGAAC-3'; the sequence of the downstream internal primer A is: 5'-TAGCTGGGCAATGGAATCCGAGGTCTCCAGAAGACCAAAGGGC-3'; the sequence of the upstream external primer A is: 5'-TCCATCTTTTGGGACCACTGT-3'; The sequence of the outer primer A is: 5'-CGACACTCTCGTCTACTCCA-3';

[0070] The volume ratio of the upstream in...

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Abstract

The invention pertains to a test technology of transgenic ingredients in food and feed, and especifically relates to a kit and a method for testing Roundup Ready transgenic Soybean and processed products thereof by using a loop-mediated isothermal amplification method. The kit comprises a loop-mediated isothermal amplification reaction solution A, a loop-mediated isothermal amplification reaction solution B, a Bst DNA polymerase C, a colour developing reagent D, and a positive control DNA E; the reaction solution A contains a reaction buffer, dNTP, magnesium sulfate, betaine, an upstream inner primer A: 5'-AGGTGGCACCTACAAATGCCATCGGCAGAGGCATCTTGAAC-3', a downstream inner primer A: 5'-TAGCTGGGCAATGGAATCCGAGGTCTCAGAAGACCAAAGGGC-3', an upstream outer primer A: 5'-TCCATCTTTGGGACCACTGT-3' and a downstream outer primer A: 5'-CGACACTCTCGTCTACTCCA-3'; and the reaction solution B contains a reaction buffer, dNTP, magnesium sulfate, betaine, an upstream inner primer B: 5'-AATCGAGCGGCGCCTCAGGCCGTAAGGAAGGCGACACC-3', a downstream inner primer B: 5'-AATGCCGCCACGGGCTCGTCGCCGATGAAGGTG-3', an upstream outer primer B: 5'-CCATGGGCGCCAGGAT-3' and a downstream outer primer B: 5'-ATCGGGCGCTTTGTGAG-3'. The method for testing the Roundup Ready transgenic Soybean and processed products thereof comprises the steps of the extraction of sample DNA, the loop-mediated isothermal amplification of cauliflower mosaic virus (CaMV) 35S promoter and Roundup Ready CP4-EPSPS gene and colour development test. The invention has the advantages of rapidness, strong specificity, high sensitivity, simple and convenient operation, good repeatability and low cost.

Description

technical field [0001] The invention relates to a method for rapid detection of transgenic products by using loop-mediated isothermal amplification technology, in particular to a method for rapid detection of glyphosate-resistant transgenic soybeans and processed products thereof. Background technique [0002] Soybean is an important oil crop and plant protein source. It is rich in protein, balanced in amino acid composition, rich in linoleic acid, lecithin, vitamin E and other nutritional factors, and is currently one of the most widely used food and feed ingredients. Since the planting of glyphosate-resistant soybeans began in 1996, glyphosate-resistant soybeans have been quickly accepted by farmers due to their advantages of greatly reducing field labor, reducing production costs, and alleviating environmental degradation, and the planting area has continued to expand. At present, glyphosate-resistant soybeans have become one of the main transgenic products in the interna...

Claims

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Application Information

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IPC IPC(8): C12Q1/68G01N21/78
Inventor 孙敏梁成珠高宏伟刘彩霞林超
Owner INSPECTION & QUARANTINE TECH CENT SHANDONG ENTRY EXIT INSPECTION & QUARANTINE BUREAU
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