Primer for testing cauliflower mosaic virus (CaMV) 35S promoter in food and feed

A cauliflower and promoter technology, applied in the field of rapid screening of transgenic products, can solve the problems of 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

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Problems solved by technology

Currently, among DNA-based detection methods, polymerase chain reaction (PCR) is mai...

Method used

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  • Primer for testing cauliflower mosaic virus (CaMV) 35S promoter in food and feed
  • Primer for testing cauliflower mosaic virus (CaMV) 35S promoter in food and feed

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Follow the procedure below for testing:

[0029] (1) DNA extraction of the tofu sample to be tested

[0030] A. Weigh 0.1g of tofu, cut it into pieces, and transfer it to a 1.5mL centrifuge tube. Add 600 μL of preheated lysis buffer, mix gently, and keep warm in a water bath at 65°C for 10 minutes;

[0031] B. Add an equal volume of 600 μL of phenol / chloroform into the tube, mix well by inverting up and down, and extract for 2 minutes;

[0032] C. Centrifuge at 12000g for 5min, draw the supernatant into a new centrifuge tube, add the same volume of sedimentation solution as the supernatant, mix well, leave at room temperature for 10min, centrifuge at 12000g for 5min, remove the supernatant, and keep the precipitate;

[0033] D. Add 60 μL of RNase to the precipitate, place it at 37°C for 2 minutes, mix it well with a pipette tip, dissolve the precipitate at 37°C, add 300 μL of buffer solution after 5 minutes, and mix up and down 10 times;

[0034] E. Take out the spin...

Embodiment 2

[0047] Follow the procedure below for testing:

[0048] (1) Extraction of the DNA of the rice sample to be tested

[0049] A. Take an appropriate amount of rice, grind it, weigh 0.1g and transfer it to a 1.5mL centrifuge tube. Add 600 μL of preheated lysis buffer, mix gently, and keep warm in a water bath at 65°C for 10 minutes;

[0050] B. Add an equal volume of 600 μL of phenol / chloroform into the tube, mix well by inverting up and down, and extract for 2 minutes;

[0051] C. Centrifuge at 12000g for 5min, draw the supernatant into a new centrifuge tube, add the same volume of sedimentation solution as the supernatant, mix well, leave at room temperature for 10min, centrifuge at 12000g for 5min, remove the supernatant, and keep the precipitate;

[0052] D. Add 60 μL of RNase to the precipitate, place it at 37°C for 2 minutes, mix it well with a pipette tip, dissolve the precipitate at 37°C, add 300 μL of buffer solution after 5 minutes, and mix up and down 10 times;

[00...

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Abstract

The invention pertains to a test technology of transgenic ingredients in food and feed, and specifically relates to a primer pair used for testing a common promoter used for transgenic plants in food and feed, namely, cauliflower mosaic virus (CaMV) 35S promoter. The invention designs a pair of specific primers, aiming at the cauliflower mosaic virus (CaMV) 35S promoter and according to the genetic conserved sequence thereof. The primer pair is used and a loop-mediated isothermal amplification technology is adopted for rapidly, sensitively and specifically testing the CaMV 35S promoter, thus screening whether the products contain transgenic plant ingredients. The primer can also be provided in kit and together with other reagents and used for nucleic acid amplification. The method has simple and convenient operation and good repeatability.

Description

technical field [0001] The invention relates to a method for rapid screening of transgenic products using loop-mediated isothermal amplification technology, in particular to a primer sequence used for rapid detection of cauliflower mottle virus CaMV 35S promoter, which is commonly used in transgenic plants. Background technique [0002] Since the first transgenic crop came out in 1983, transgenic technology has been rapidly applied and developed in various fields of agriculture. The variety of genetically modified crops in the world is increasing continuously, and the planting area has reached 690 million hectares. The use of transgenic technology can precisely improve and improve the quality and quantity of crops. Compared with traditional crops, genetically modified crops have significantly improved yield, stress resistance (including disease resistance, insect resistance, frost resistance, herbicide resistance) and nutritional quality, which can greatly reduce production...

Claims

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Application Information

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IPC IPC(8): C12Q1/68G01N21/78C12N15/11
Inventor 孙敏徐彪林超高宏伟孙雯娴刘彩霞
Owner INSPECTION & QUARANTINE TECH CENT SHANDONG ENTRY EXIT INSPECTION & QUARANTINE BUREAU
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