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Primers, probes and methods for accurate detection of specific quantitative PCR for transgenic maize mir604 transformation events

A technology for transgenic corn and transformation events, applied in the field of quantitative analysis, can solve the problems of no amplification detection of transgenic corn MIR604 transformation events, etc., and achieve the effects of protecting the right to know and the right to choose, high amplification efficiency, and high accuracy

Inactive Publication Date: 2016-06-22
四川省农业科学院分析测试中心
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Problems solved by technology

[0003] At present, the detection technology of transgenic corn MIR604 mainly focuses on common qualitative PCR analysis methods and standards, and there is no accurate quantitative PCR detection technology for amplifying and detecting a specific site (gene sequence) of the transformation event of transgenic corn MIR604 and its products

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  • Primers, probes and methods for accurate detection of specific quantitative PCR for transgenic maize mir604 transformation events
  • Primers, probes and methods for accurate detection of specific quantitative PCR for transgenic maize mir604 transformation events

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Experimental program
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Embodiment

[0035] The method for accurate detection of transgenic maize MIR604 transformation event-specific quantitative PCR mainly includes the following steps:

[0036] (1) Synthesize the following primers and fluorescent probes used in conjunction with the primers,

[0037] Sequence of upstream primer, MIR604eveF: 5'-GCCGTATCCGCAATGTGTTAT-3'

[0038] Downstream primer sequence, MIR604eveR: 5'-CGACGCTTGCGCGAG-3'

[0039] Fluorescent probe sequence, MIR604eveP: 5'-FAM-AGAGTTGGTGGTACGGGTA-TAMRA-3'.

[0040] The synthesis concentrations of the primers and fluorescent probes in this example were both 10 μmol / l.

[0041] The nucleotide sequences of the above primers and fluorescent probes are designed for a specific site specific to the transformation event of transgenic maize MIR604 and its products, that is, the target gene and the flanking site of the recipient maize genome; through this design, the transgene can be accurately detected Transformation events of MIR604 in maize.

[00...

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Abstract

The invention belongs to the field of biotechnology, relates to a quantitative gene analysis method and in particular relates to a transgenic maize MIR604 transformation event specific quantitative PCR (polymerase chain reaction) accurate detection method. A PCR reaction system is prepared from a designed specific forward primer sequence MIR604eveF, a reverse primer sequence MIR604eveR, a fluorescence probe sequence MIR604eveP, a DNA diluent of MIR604, TaqmanMastermix(2X) and water, and then quantitative PCR detection is performed. A Taqman quantitative PCR detection technology with high amplification efficiency and high accuracy is mainly established, and the method is applied to supervision and inspection of domestic genetically modified organisms and products in agriculture, inspection of import and export genetically modified organisms and products and detection of inner-enterprise import raw materials containing transgenic MIR604-strain biological components.

Description

technical field [0001] The invention belongs to the field of biotechnology and relates to a quantitative analysis method of genes. Background technique [0002] Many countries in the world implement limited labeling and importation of genetically modified products, but there is no specific labeling threshold for genetically modified products in my country. In order to break the technical barriers to the trade of genetically modified products set up by the European Union and other countries and regions, at the same time make up for and improve the quantitative detection technology system of genetically modified organisms and products in my country, and better protect consumers' right to know and to choose genetically modified products, a new type of genetically modified The accurate detection method of specific quantitative PCR for maize MIR604 transformation event has become necessary. [0003] At present, the detection technology of transgenic corn MIR604 mainly focuses on ...

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6851C12Q2531/113C12Q2561/101
Inventor 常丽娟宋君雷绍荣郭灵安刘勇王东尹全刘文娟张富丽
Owner 四川省农业科学院分析测试中心
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