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Method for extracting DNA for trangene ingredient detection from plant source edible oil

A technology of genetically modified ingredients and edible oil, applied in the field of nucleic acid extraction, can solve problems such as unstable extraction, and achieve the effect of good stability and high efficiency

Inactive Publication Date: 2010-12-29
TIANGEN BIOTECH BEIJING
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Our invention can overcome the above-mentioned defects of unstable extraction

Method used

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  • Method for extracting DNA for trangene ingredient detection from plant source edible oil
  • Method for extracting DNA for trangene ingredient detection from plant source edible oil
  • Method for extracting DNA for trangene ingredient detection from plant source edible oil

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0065] Using commercially available Arowana transgenic soybean oil and Fulinmen transgenic soybean oil as materials, the following method was used to extract DNA,

[0066] (1) Take 30ml of soybean oil, add an equal volume of n-hexane, and stir on a magnetic stirrer for 2-3 hours.

[0067] (2) Take 10 ml of the pretreated solution, add 5 ml of extraction buffer, and vortex for 1 minute. Let stand at room temperature for 10 minutes.

[0068] (3) Add 2ml of sodium acetate buffer, mix thoroughly, and vortex for 1 minute. Let stand at room temperature for 10 minutes.

[0069] (4) Centrifuge at 10,000 rpm for 10 minutes, and transfer the lower aqueous phase to a new centrifuge tube.

[0070] (5) Add 10 μl of Carrier RNA and 0.7 times volume of isopropanol to the aqueous phase solution, and mix thoroughly. (For example, 5ml of aqueous phase solution plus 3.5ml of isopropanol), centrifuge at 10,000rpm for 10 minutes, discard the supernatant, and keep the precipitate.

[0071] (6)...

Embodiment 2

[0079] Material, step are with embodiment 1.

[0080] Soybean seed DNA extracted with Tiangen DP321 kit was used as a control. Using the specific primers of the transgenic soybean promoter CaMV35S, and using the DNA extracted by this method as a template, check whether the extracted DNA meets the requirements for the detection of genetically modified components by PCR amplification. The results are shown in figure 1 .

[0081] Depend on figure 2 It can be seen that the DNA extracted from Arowana transgenic soybean oil and Fulinmen transgenic soybean oil was amplified by PCR using CaMV35S promoter-specific primers, and the amplified fragments obtained were the same size as the amplified fragments of control soybean seeds. There is no amplification in the control, so the DNA extracted by this method can fully meet the requirements for the detection of genetically modified components.

Embodiment 3

[0083] Using the commercially available Emerald Rapeseed Oil as the material, the following method was used to extract DNA,

[0084] (1) Take 30ml of rapeseed oil, add an equal volume of n-hexane, and stir on a magnetic stirrer for 2-3 hours.

[0085] All the other steps are the same as in Example 1.

[0086] Rapeseed DNA extracted with the Tiangen DP321 kit was used as a control. Using the specific primers of rapeseed endogenous gene PE3PEPcase, using the DNA extracted by this method as a template, check whether the extracted DNA meets the requirements for the detection of genetically modified components by PCR amplification. The results are shown in figure 1 .

[0087] Depend on figure 2 It can be seen that the DNA extracted from rapeseed oil, after using the specific primers of PE3PEPcase gene for PCR amplification, the amplified fragment obtained is consistent with the size of the amplified fragment of the control rapeseed, while the clear water control has no amplific...

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Abstract

The present invention relates to a method for extracting a DNA used for the detection of genetically modified components from edible vegetable oil. The method has the following characteristics: before the extraction of the DNA, the edible vegetable oil is emulsified by organic emulsifier for a long time; the extremely low-content DNA is sufficiently dissociated out, so that the first step of successful extraction is fulfilled; an independently developed unique buffer system (extraction buffer solution 00mM Tris.HCl, pH8.0; 20mM EDTA; 500mM NaCl; 1.5 percent of SDS; 4 percent of PVP) is utilized to sufficiently release and aggregate the DNA, which is different from the conventional CTAB cracking and phenol / chloroform extraction; in carrier RNA coprecipitated catalyst, isopropanol precipitation is carried out twice; the solution is transferred from a big centrifugal tube to a small centrifugal tube; the extremely low-content DNA obtained from the edible vegetable oil to the max is dissolved in 40Wu dissolving buffer solution; 9Wu1 is taken out to serve as a template which is used for the PCR reaction system of 20Wu1. The method, which has the advantages of rapidity, convenience, safety, high efficiency and good stability, is applicable to the detection of the genetically modified components of edible vegetable oil.

Description

technical field [0001] The invention belongs to the technology in the field of nucleic acid extraction, and in particular relates to a nucleic acid extraction method for detection of genetically modified components of plant source edible oil. Background technique [0002] Since the development of GMOs and GMO products in the 1970s, GMO products have been developed at a rapid pace. According to the statistics of international authoritative organizations, there are nearly 10,000 kinds of genetically modified foods and food ingredients processed and produced from genetically modified crops in the world. Among them, soybean and rapeseed, which are used as raw materials for edible oil processing, occupy a large proportion. For example, soybean accounts for 63% of the total planting area of ​​transgenic crops in the world, and rapeseed accounts for 5%. Due to the high oil yield of these genetically modified oil crops and the low cost of oil production, developing countries have i...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/10C12Q1/68
Inventor 柯海燕
Owner TIANGEN BIOTECH BEIJING