Visualized detection method of BAR transgenic crops

A detection method and crop technology, applied in the field of genetic engineering, can solve the problem of high detection cost of fluorescent probes, achieve the effects of cheap reagents, lower detection costs, and convenient synthesis

Inactive Publication Date: 2012-11-21
CHENGDU INST OF BIOLOGY CHINESE ACAD OF S
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Among them, the first type of method is widely used. The detection method based on nucleic acid is mainly based on the detection method of fluorescent quantitative PCR. Fluorescent quantitative PCR has many adv

Method used

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  • Visualized detection method of BAR transgenic crops
  • Visualized detection method of BAR transgenic crops
  • Visualized detection method of BAR transgenic crops

Examples

Experimental program
Comparison scheme
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Example Embodiment

[0020] Example 1: Detection of BAR gene in rice

[0021] 1. Extraction of rice genomic DNA;

[0022] The extraction of rice genomic DNA is by the improved CTAB method. ①A proper amount of rice leaves or seeds are ground into fine powder in a mortar under the protection of liquid nitrogen. Take a small amount of the powder into a 1.5mL EP tube, add 700μl of CTAB extraction buffer preheated at 65℃, shake well and mix well, and incubate at 65℃ for 2h ②After the sample is cooled to room temperature, add 600μl of chloroform-isoamyl alcohol (24:1) and mix by inversion for 5 minutes; ③Centrifuge at 7500r.min-1 for 10 minutes, transfer the supernatant to a new EP tube; ④Add, etc. Volume of isopropanol pre-cooled at -20°C, mix well, and place at 20°C for 30 minutes; ⑤Centrifuge at 10,000 r.min-1 for 15 minutes, discard the supernatant; ⑥ Wash the precipitation with 70% ethanol and anhydrous ethanol once, Centrifuge at 10,000r.min-1 for 3 minutes, discard the supernatant, evaporate the eth...

Example Embodiment

[0033] Example 2: Detection of BAR gene in cotton

[0034] 1. Extraction of cotton genomic DNA;

[0035] The genomic DNA of cotton is extracted by a modified CTAB method. ① An appropriate amount of cotton leaves or seeds are ground into a fine powder in a mortar under the protection of liquid nitrogen. Take a small amount of the powder into a 1.5mL EP tube, add 700μl of CTAB extraction buffer preheated at 65℃, shake well and mix well, and incubate at 65℃ for 2h ②After the sample is cooled to room temperature, add 600μl of chloroform-isoamyl alcohol (24:1) and mix by inversion for 5 minutes; ③Centrifuge at 7500r.min-1 for 10 minutes, transfer the supernatant to a new EP tube; ④Add, etc. Volume of isopropanol pre-cooled at -20°C, mix well, and place at 20°C for 30 minutes; ⑤Centrifuge at 10,000 r.min-1 for 15 minutes, discard the supernatant; ⑥ Wash the precipitation with 70% ethanol and anhydrous ethanol once, Centrifuge at 10,000r.min-1 for 3 minutes, discard the supernatant, eva...

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Abstract

The invention belongs to the technical field of genetic engineering and relates to a detection technology of transgenic crops, in particular to a visualized detection method of BAR transgenic crops. The method comprises steps as follows: adding DNAzyme-marked specific probes for detecting the BAR gene into a PCR (polymerase chain reaction) system, amplifying the BAR gene by forward and reverse primers, releasing the DNAzyme markers from the probes by exonuclease activity of nucleic acid polymerase 5'-3', and detecting the markers released from the PCR product to judge whether the sample is a transgenic sample or not. The method provided by the invention has the characteristics of being efficient and convenient and low in cost and the like.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and relates to a detection technology of transgenic crops, in particular to a visual detection method of transgenic BAR gene crops. Background technique [0002] Genetically modified crops are bred by introducing genes from other organisms or species by using tissue culture technology and genetic recombination technology. Such crops often have characteristics such as resistance to diseases and insect pests, resistance to herbicides, or higher nutrition than non-genetically modified crops. At present, there are more than 20 kinds of genetically modified plants in China, among which genetically modified soybeans, potatoes, corn, tobacco, peanuts, spinach, sweet peppers, wheat, etc. have undergone field tests, genetically modified cotton has been used on a large scale, and foreign genetically modified soybeans have also entered my country in large quantities. Although genetically modifie...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 唐卓余懋群陈蓉龙海
Owner CHENGDU INST OF BIOLOGY CHINESE ACAD OF S
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