Primer and probe for fluorescence quantitative PCR of phomopsis amydalina TaqMan

A Phomopsis and fluorescence quantitative technology, which is applied in the direction of microorganisms, microorganism-based methods, biochemical equipment and methods, etc., can solve problems such as the reliability of detection results, and achieve reliable and easy-to-operate detection results

Inactive Publication Date: 2016-06-15
WUXI ENTRY EXIT INSPECTION & QUARANTINE BUREAU PEOPLES REPUBLIC OF CHINA
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Problems solved by technology

The molecular detection technology developed in the early stage lacks the support of genomics, and the molecular detect

Method used

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  • Primer and probe for fluorescence quantitative PCR of phomopsis amydalina TaqMan
  • Primer and probe for fluorescence quantitative PCR of phomopsis amydalina TaqMan
  • Primer and probe for fluorescence quantitative PCR of phomopsis amydalina TaqMan

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Embodiment 2

[0053] Embodiment 2: Actual sample detection

[0054] The 50 samples collected in the field were detected by isolation culture method, conventional PCR method and fluorescent quantitative PCR method. The results showed that in the TaqMan fluorescent quantitative PCR, 43 samples showed obvious amplification curves, with a detection rate of 86%; in conventional PCR, 39 samples were positive, with a detection rate of 78%; while only 36 samples in the culture method Phomopsis amygdala was successfully isolated, and the detection rate was 72% (as shown in Table 4). Therefore, it can be considered that the sensitivity of fluorescent quantitative PCR detection samples is higher than that of culture method and conventional PCR. At the same time, the separation and culture method has a long experimental cycle, the whole detection process needs 72h, and the workload is large, and it is susceptible to the interference of saprophytic bacteria to cause deviations in the experimental result...

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Abstract

The invention relates to a primer and a probe for a fluorescence quantitative PCR of phomopsis amydalina TaqMan. The primer is characterized by comprising a primer pair formed by an upstream primer histone H3-F shown as the SEQ ID No:1 and a downstream primer histone H3-R shown as the SEQ ID No:1. The probe used for detecting phomopsis amydalina is a probe histone H3-P shown as the SEQ ID No:1. The end 5' of the probe histone H3-P marks a fluorescence reporter group FAM, and the end 3' of the probe histone H3-P marks a fluorescence quenching group TAMRA. A method for detecting phomopsis amydalina is characterized by comprising the steps that 1, the upstream primer histone H3-F, the downstream primer histone H3-R and the probe histone H3-P are synthesized, and the probe histone H3-P is marked; 2, genome DNA of a sample to be detected is extracted, and amplification is performed with the upstream primer histone H3-F and the downstream primer histone H3-R to obtain an amplification product; 3, a fluorescence quantitative PCR detection system is adopted for detecting the sample to be detected. The primer and the probe are suitable for fluorescence quantitative PCR detection of phomopsis amydalina TaqMan, and rapidity, sensitivity, simplicity and convenience are achieved.

Description

technical field [0001] The invention relates to Phomopsis amygdala TaqMan fluorescent quantitative PCR primers and probes, which belong to the field of biotechnology and are suitable for use in port inspection and quarantine, agricultural production, plant protection and other departments. Background technique [0002] Phomopsis amygdalis (asexual state: Phomopsisamygdali, sexual state: Diaportheamygdali) is an important plant pathogen, which is pathogenic to many Rosaceae plants such as peach, red leaf plum, flower plum, pear and apricot, but in peach Peach constriction canker caused on trees is particularly serious. It often manifests as the formation of ring-shaped brown disease spots at the base of the new shoots and twigs of peach trees, causing the leaves above the diseased parts of the branches to wither and fall off quickly, and the disease spreads and expands rapidly, which can cause 20-50% yield loss, and seriously cause the whole peach tree to rot. Strains die. ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12Q1/04C12N15/11C12R1/645
CPCC12Q1/6851C12Q1/6895C12Q2531/113C12Q2561/101C12Q2545/113
Inventor 张华胡白石
Owner WUXI ENTRY EXIT INSPECTION & QUARANTINE BUREAU PEOPLES REPUBLIC OF CHINA
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