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Real-time fluorescence PCR (polymerase chain reaction) detection method based on ITS (internal transcribed spacer) gene for Opogona sacchari

A real-time fluorescence, detection method technology, applied in fluorescence/phosphorescence, microbial determination/inspection, biochemical equipment and methods, etc. The effect of public health safety, accurate and stable detection, and low detection limit

Inactive Publication Date: 2012-10-31
中华人民共和国珠海出入境检验检疫局 +1
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Problems solved by technology

This method is difficult to operate, it is difficult to obtain enough COI genes, and subsequent analysis will consume a lot of time

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  • Real-time fluorescence PCR (polymerase chain reaction) detection method based on ITS (internal transcribed spacer) gene for Opogona sacchari

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Embodiment Construction

[0014] The gene encoding ribosomal RNA (rDNA) is the most conserved structural gene in eukaryotes. It is arranged in an adaptable repeat, each repeat including three coding regions (respectively encoding 18S, 5.8S and 28S RNA genes), encoding The regions are separated by two ITS (Internal Transcribed Spacers), and the repeats are separated by an ETS (External Transcribed Spacer) and an IGS (Integenic Spacer) respectively. Since rDNA has hundreds of copies in the genome (increased Sensitivity of detection) and the stability of ITS and IGS regions within species and differences between species, making it an important object of genetic variation research in many organisms, and also a hot spot in insect genetic variation, phylogenetic and molecular identification research in recent years , existing studies have shown that: the ITS regions of many insects in the same group and different species are highly conserved in the length of the nucleotide sequence, and there is no obvious di...

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Abstract

The invention discloses a real-time fluorescence PCR (polymerase chain reaction) detection method based on ITS (internal transcribed spacer) gene for Opogona sacchari, comprising the following steps of: extracting the DNA of a sample to be detected, preparing the DNA into a template, performing real-time fluorescence PCR by an upstream primer, a downstream primer and a probe, and determining the variety of the sample according to the Ct value. The method is quick, stable and reliable, has high sensitivity, and is suitable for detecting different stages of Opogona sacchari. The method lays a foundation for classification on the molecular level and quick identification of allied species for Opogona (Lepidoptera: Tineidae) in China. The research results can be directly applied to quarantine and identification of Opogona sacchari in plant passed in and out quarantine, thus having great importance for protecting production safety in agriculture and forestry in China and maintaining the ecological balance.

Description

technical field [0001] The invention relates to a molecular identification method of biological species, in particular to a real-time fluorescent PCR detection method of flat moth sucrose moth based on ITS gene. Background technique [0002] Cane flat moth Opogona sacchari (Bojer) is a new quarantine pest in my country. This insect is also listed as a quarantine insect in regions such as the European Plant Protection Organization (EPPO), the Asia-Pacific Plant Protection Organization (APPO) and the North American Plant Protection Organization (NAPPO). . It was first discovered in Beijing garden plants in 1997. The insect has a wide range of hosts, 24 families and 46 species have been reported; it can harm bananas, sugarcane, Brazil wood Dracaena fragrana (L.), Fortune Tree Pachira macrocarpa And other garden plants. According to reports, the flat moth was first discovered in Mauritius, and then found in nearby Madagascar and the Seychelles Islands in the north. Soon...

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68G01N21/64
Inventor 廖力徐淼锋张卫东黄国华
Owner 中华人民共和国珠海出入境检验检疫局
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