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A common primer and detection method for identification of potato scab pathogen species

A technology for potato scab and scab bacteria, applied in the direction of microorganism-based methods, biochemical equipment and methods, and microbial measurement/inspection, can solve the problems of low identification efficiency, low identification resolution, and poor accuracy, and achieve identification High efficiency, high resolution and good accuracy

Active Publication Date: 2020-09-08
黑龙江省农业科学院植物脱毒苗木研究所
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to solve the problem of low identification resolution, low identification efficiency and poor accuracy of the pathogenic bacteria of potato scab

Method used

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  • A common primer and detection method for identification of potato scab pathogen species
  • A common primer and detection method for identification of potato scab pathogen species
  • A common primer and detection method for identification of potato scab pathogen species

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specific Embodiment approach 1

[0025] Specific embodiment one: a kind of potato scab pathogen species identification universal primer of the present embodiment, this primer pair is as follows:

[0026] gyrBF2: 5'-TCTGCACGGYGTSGGYGTCTC-3'

[0027] gyrBR2: 5'-TTGCGGGAGTTGAGGTAC-3'.

[0028] The primer sequence of this embodiment is shown in the sequence table Seq ID No: 8, and the sequence table Seq ID No: 9.

specific Embodiment approach 2

[0029] Specific embodiment two: a kind of detection method of identification potato scab bacterium kind of the present embodiment, it is to carry out according to the following steps:

[0030] 1. Extract the genomic DNA of the sample to be tested;

[0031] 2. Using the primers of the specific embodiment 1 as identification primers, the DNA extracted in step 1 is used as a template to carry out PCR amplification reaction;

[0032] 3. PCR products were subjected to 1% agarose gel electrophoresis;

[0033] 4. Analyze the amplified slice: there is an amplified band at the 400bp position, indicating that the PCR product of the sample can be sequenced, and then sequenced;

[0034] 5. Comparing the sequenced sequence with the data in the gene database. After the comparison is successful, the detection for identifying the species of potato scab bacteria is completed.

specific Embodiment approach 3

[0035] Specific embodiment three: the difference between this embodiment and specific embodiment two is: the PCR system of PCR amplification is as follows:

[0036]

[0037] PCR amplification conditions: pre-denaturation at 95°C for 5 min; denaturation at 95°C for 45 s, annealing at 50-65°C for 45 s, extension at 72°C for 90 s, 35 cycles, extension at 72°C for 10 min, and storage at 4°C.

[0038] Others are the same as in the second embodiment.

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Abstract

The invention provides a universal primer for identifying pathogenic species of potato scabs and a detection method, which relates to a detection primer and a PCR (polymerase chain reaction) amplification method. The invention aims at solving the problems of low identification resolution ratio, low identification efficiency and poor identification accuracy of the pathogenic species of the potato scabs. The sequences of the primer are gyrBF2:5'-TCTGCACGGYGTSGGYGTCTC-3' and gyrBR2:5'-TTGCGGGAGTTGAGGTAC-3'. The detection method comprises the following steps: extracting genomic DNA of a bacterial strain, performing PCR amplification, sequencing products with fragment size of 400bp, comparing a sequencing result and a gene database and finally determining the pathogenic spices. The universal primer has comparatively high identification resolution ratio between spices, high identification efficiency and high repeatability. A detected sample can be pure bacterial strain DNA, and potato tuber tissue DNA after being infected by a single pathogenic bacterium serves as a template for PCR amplification and sequencing.

Description

technical field [0001] The invention relates to a detection primer and a PCR amplification method thereof. Background technique [0002] Potato scab (Potato common scab), a soil-borne disease that affects the appearance, grade and quality of potatoes, is widely distributed and is a serious world-class disease. Potato scab mainly harms tubers. Pathogens invade through the lenticels. Brown spots appear on the tuber epidermis at the beginning, and then gradually expand. The tissue around the infection point is necrotic. The severity of the disease varies with varieties, plots, and years, and the size and depth of lesions also vary with the variety of pathogenic bacteria, the susceptibility of varieties, and the environmental conditions. In severe cases, the lesions are contiguous, seriously reducing the appearance quality of the tubers and affecting their commodity value and processing value. Economic losses due to scab range from US$90-102 per hectare in Canada and US$153-17...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/689C12Q1/686C12Q1/04C12N15/11C12R1/465
CPCC12Q1/686C12Q1/689C12Q2565/125
Inventor 魏琪闵凡祥王晓丹高云飞杨帅董学志王文重张抒吕典秋宿飞飞张静华李辉
Owner 黑龙江省农业科学院植物脱毒苗木研究所
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