Real-time fluorescent RT-PCR detection method, primer and probe for oat mosaic virus and kit

A technology of RT-PCR and mosaic virus, which is applied in the direction of microorganism-based methods, biochemical equipment and methods, and microorganism measurement/inspection, to achieve the effects of reducing human error, improving sensitivity, and quantitative accuracy

Pending Publication Date: 2017-08-15
珠海出入境检验检疫局检验检疫技术中心
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AI Technical Summary

Problems solved by technology

However, so far, there is a lack of a real-time fluorescent quantitative RT-PCR detection method with strong specificity and high sensitivity for oat mosaic virus

Method used

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  • Real-time fluorescent RT-PCR detection method, primer and probe for oat mosaic virus and kit
  • Real-time fluorescent RT-PCR detection method, primer and probe for oat mosaic virus and kit
  • Real-time fluorescent RT-PCR detection method, primer and probe for oat mosaic virus and kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment

[0032] 1. Select the test material. The virus samples used for the test are shown in Table 1.

[0033] Table 1 Virus samples for testing

[0034] Chinese name English name oat mosaic virus Oat mosaic virus, OMV barley yellow mosaic virus Barley yellow mosaic virus, BYMV barley and sexual mosaic virus Barley mild mosaic virus, BaMMV wheat yellow mosaic virus Wheat yellow mosaic virus, WYMV Wheat shuttle streak mosaic virus Wheat spindle streak mosaic virus, WSSMV rice necrotic mosaic virus Rice necrosis mosaic virus, RNMV

[0035] The main reagents and instruments used in this example are as follows: fluorescent PCR amplification reagents were provided by Zhuhai Baorui Biotechnology Co., Ltd.; RNA extraction kits, reverse transcription kits, primers and probes were provided by Bao Bioengineering (Dalian) Co., Ltd. Provided by the company; other reagents are in accordance with the regulations of ordinary molecular biolog...

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Abstract

The invention relates to the field of molecular diagnosis of biological species and discloses a real-time fluorescent RT-PCR detection method, primer and probe for an oat mosaic virus and a kit. The detection method comprises the following steps: (S1) extracting total RNA of plant tissue from a to-be-detected sample; (S2) introducing a downstream primer OMV-R1 for synthesis of a reverse transcription template cDNA by employing the total RNA as a template; (S3) putting an upstream primer OMV-F1, the downstream primer OMV-R1 and a fluorescent probe OMV-P into a fluorescent quantitative PCR instrument for real-time quantitative fluorescent PCR amplification reaction; and (S4) reading a Ct value to judge the detection result through the fluorescent PCR instrument. A specific primer and a fluorescent probe are designed for a quarantine pest oat mosaic virus; a method for quickly detecting OMV through real-time quantitative fluorescent RT-PCR is built; the method has the advantages of being simple in detection method (closed tube in the whole process and free of electrophoresis detection), high in speed, quantitative, accurate, high in specificity and high in sensitivity, and is suitable for various inspection and quarantine institutions in China for quarantine identification of the quarantine pest oat mosaic virus.

Description

technical field [0001] The invention relates to the field of molecular diagnosis of biological species, in particular to a real-time fluorescent RT-PCR detection method for oat mosaic virus, primers, probes and a kit. Background technique [0002] Oat mosaic virus (OMV) belongs to the Potyviridae family and belongs to the genus Bymovirus. It is linear, often curved, and has no envelope. It has an obvious morphological length, about 600~ 750nm, 12-14nm wide. OMV mainly infects plants of the genus Avena, causing disease of oats sown in autumn, and can cause a 39-60% yield reduction. The spread of OMV mainly depends on the persistent transmission of Polymyxa graminearum, and the long-distance spread is spread with the vegetative material of susceptible plants. At present, OMV is mainly distributed in the United States, Canada, New Zealand, the United Kingdom, France, Ireland and other countries, and has not yet occurred in my country. In 2007, the Ministry of Agriculture's of...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/68C12N15/11C12R1/94
CPCC12Q1/686C12Q1/701C12Q2521/107C12Q2563/107
Inventor 张卫东黄永辉权永兵廖力徐淼锋迟远丽
Owner 珠海出入境检验检疫局检验检疫技术中心
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