A fluorescent detection kit for 27 plant transgenic loci complex PCR amplification

A detection kit and genetically modified technology, which is applied in the determination/inspection of microorganisms, biochemical equipment and methods, etc., can solve the problems of insufficient accuracy and sensitivity, and achieve accurate results

CN104593504BActive Publication Date: 2017-01-18AGCU SCIENTECH +1
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Patent Information

Authority / Receiving Office
CN · China
Patent Type
Patents(China)
Current Assignee / Owner
Publication Date
2017-01-18

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Abstract

The invention discloses a composite PCR (polymerase chain reaction) amplification fluorescence detection kit for 27 plant transgenic loci. The kit comprises a mixture of a PCR buffer solution, MgCl2, dNTPs and bovine serum albumin, a hot-start rapid Taq enzyme, 27 pairs of primer mixtures and ultra-pure water. Whether the transgenic loci contained in the kit exist or not can be detected only by carrying out nucleic acid amplification on a plant genome DNA sample for about 80 minutes; and the detection kit is suitable for identifying transgenic ingredients or strains of corn, soybean, paddy and oilseed rape, and is a rapid, simple, economic and efficient transgenic ingredient detection kit.
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Description

technical field

[0001] The invention relates to a 27 plant transgenic site complex PCR amplification fluorescent detection kit, and the system can be used for identification of transgenic components or strains of corn, soybean, rice and rapeseed. Background technique

[0002] Transgenic technology refers to the introduction of artificially isolated and modified genes into the genome of organisms, resulting in the modification of heritable traits of organisms, and obtaining genetically modified organisms (GMOs). Genetically modified crops have characteristics such as insect resistance, disease resistance, and high yield, and are considered to lead to the second green revolution.

[0003] So far, at least 28 countries around the world are planting genetically modified crops, the planting area has exceeded 170 million hectares, and the sales of genetically modified seeds reached 15 billion US dollars in 2012. Various countries and regions have strict testing standards and requ...

Examples

Embodiment 1

[0047] 1. Extraction of sample DNA

[0048] Using the CTAB method to extract the sample seed powder DNA, the specific steps are as follows:

[0049] (1) Preheat the CTAB solution at 60°C for 30 minutes;

[0050](2) Weigh 0.2g of seed powder into a 1.5mL centrifuge tube, add 700ul of CTAB solution, shake, and bathe in 60°C water bath for 30 minutes;

[0051] (3) Add 700 μL of chloroform:isoamyl alcohol (24:1), mix upside down, and centrifuge at 12,000 rpm for 10 minutes at room temperature;

[0052] (4) Aspirate the supernatant, add an equal volume of chloroform:isoamyl alcohol (24:1), mix upside down, and centrifuge at 12,000 rpm for 10 minutes at room temperature;

[0053] (5) Aspirate the supernatant, add 0.8 times the volume of isopropanol, mix gently, and ice bath for 20 minutes;

[0054] (6) Centrifuge at 12000rpm for 10 minutes, discard the supernatant, and dry the water;

[0055] (7) Add 500 μL of 70% ethanol, pipette the precipitate, centrifuge at 12000 rpm for 1 m...