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Tomato seed-borne pathogen multiple PCR detection kit and tomato seed-borne pathogen multiple PCR detection method

A detection kit and detection method technology, applied in biochemical equipment and methods, microbial determination/inspection, etc., can solve problems such as no reports of multiple PCR detection methods, achieve efficient and rapid detection, avoid repeated operations, and stabilize good effect

Active Publication Date: 2014-12-24
兰州海关技术中心
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, PSTVd, PepMV and TRSV have established a single RT-PCR molecular detection technology, but there are no reports on the multiple PCR detection methods of these three important species-borne pathogens

Method used

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  • Tomato seed-borne pathogen multiple PCR detection kit and tomato seed-borne pathogen multiple PCR detection method
  • Tomato seed-borne pathogen multiple PCR detection kit and tomato seed-borne pathogen multiple PCR detection method
  • Tomato seed-borne pathogen multiple PCR detection kit and tomato seed-borne pathogen multiple PCR detection method

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Effect test

Embodiment 1

[0039] Embodiment 1: A multiplex PCR detection kit for tomato seed-borne pathogens. The tomato seed-borne pathogen is potato spindle tuber viroid, pineapple mosaic virus or tobacco ringspot virus, which includes the following components:

[0040] RT-PCR buffer, 2×RT-PCR buffer, 250 μL; RT-PCR amplification primers are potato spindle tuber viroid PSTVd ​​upstream primers, the concentration is 10 μmol / L, 30 μL; potato spindle tuber viroid PSTVd ​​downstream primers, the concentration is 10 μmol / L, 30 μL; PepMV upstream primer, the concentration is 10 μmol / L, 30 μL; PepMV downstream primer, the concentration is 10 μmol / L, 30 μL; Tobacco ringspot virus TRSV upstream primer, the concentration is 10 μmol Tobacco ringspot virus TRSV downstream primer, concentration 10 μmol / L, 30 μL; Reverse transcriptase and DNA polymerase Mix, 20 μL; RNase-free deionized water, 1 mL; Positive control, potato spindle tubers 60 μL each of positive controls with no infectivity of virus PSTVd, pineappl...

Embodiment 2

[0048] Embodiment 2: A multiplex PCR detection kit for tomato seed-borne pathogens, tomato seed-borne pathogens are potato spindle tuber viroid, pineapple mosaic virus or tobacco ringspot virus, which includes the following components:

[0049] RT-PCR buffer, 2×RT-PCR buffer, 100 μL; RT-PCR amplification primers are potato spindle tuber viroid PSTVd ​​upstream primers, the concentration is 10 μmol / L, 10 μL; potato spindle tuber viroid PSTVd ​​downstream primers, the concentration is 10 μmol / L, 10 μL; PepMV upstream primer, the concentration is 10 μmol / L, 10 μL; PepMV downstream primer, the concentration is 10 μmol / L, 10 μL; Tobacco ringspot virus TRSV upstream primer, the concentration is 10 μmol Tobacco ringspot virus TRSV downstream primer, 10 μmol / L, 10 μL; Reverse transcriptase and DNA polymerase Mix, 20 μL; RNase-free deionized water, 100 μL; Positive control, potato spindle tubers 20 μL each of positive controls with no infectivity of virus PSTVd, pineapple mosaic virus...

Embodiment 3

[0057] Embodiment 3: A multiplex PCR detection kit for tomato seed-borne pathogens, tomato seed-borne pathogens are potato spindle tuber viroid, pineapple mosaic virus or tobacco ringspot virus, which includes the following components:

[0058] RT-PCR buffer, 2×RT-PCR buffer, 150 μL; RT-PCR amplification primers are potato spindle tuber viroid PSTVd ​​upstream primers, the concentration is 10 μmol / L, 20 μL; potato spindle tuber viroid PSTVd ​​downstream primers, the concentration is 10 μmol / L, 20 μL; PepMV upstream primer, the concentration is 10 μmol / L, 20 μL; PepMV downstream primer, the concentration is 10 μmol / L, 20 μL; Tobacco ringspot virus TRSV upstream primer, the concentration is 10 μmol / L, 20 μL; Tobacco ringspot virus TRSV downstream primer, the concentration is 10 μmol / L, 20 μL; Reverse transcriptase and DNA polymerase Mix, 20 μL; RNase-free deionized water, 50 μL; Positive control, potato spindle tubers 40 μL each of positive controls with no infectivity of viru...

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Abstract

The invention belongs to the technical field of biology, and relates to a detection method of a tomato disease. The detection method is mainly used for simultaneous detection of three important tomato seed-borne pathogens, namely a potato spindle tuber viroid (PSTVd), a pepino mosaic virus (PepMV) and a tobacco ringspot virus (TRSV). A tomato seed-borne pathogen multiple PCR (Polymerase Chain Reaction) detection kit for the tomato seed-borne pathogens, namely the PSTVd, the PepMV or the TRSV, is mainly characterized by comprising the following components: an RT-PCR (Reverse Transcription-PCR) buffer, a PSTVd upstream and downstream primer, a PepMV upstream and downstream primer, a TRSV upstream and downstream primer, a reverse transcriptase and DNA (Deoxyribose Nucleic Acid) polymerase Mix, deionized water without RNA (Ribose Nucleic Acid) enzyme, and an infection-activity-free positive control of the PSTVd, the PepMV and the TRSV. The detection method has the advantages that the three important tomato seed-borne pathogens can be detected simultaneously; the detection method is rapid and accurate; and repetitive operation of conventional PCR detection is avoided.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to a detection method for tomato diseases, which is mainly aimed at the simultaneous detection of three important tomato seed-borne pathogens, potato spindle tuber viroid, pineapple mosaic virus and tobacco ringspot virus. Background technique [0002] Tomato virus disease is an important disease caused by a variety of viruses. It is one of the most common and difficult to control diseases on tomato, and it is distributed in tomato producing areas in the world. Potato spindle tuber viroids, pineapple mosaic virus and tobacco ringspot virus can all cause tomato virus diseases. Potato spindle tuber viroid (PSTVd) is a phytosanitary pest imported into my country. It is locally distributed in Heilongjiang, Jiangsu, Hebei and Qinghai in China. Its main hosts are potatoes, tomatoes and avocados. It can also infect other plants. Plants, including capsicum, sweet potato, nightshade, nightshade or...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/70C12Q1/68
CPCC12Q1/686C12Q1/70C12Q2537/143C12Q2531/113
Inventor 文朝慧王军平周小平李儒刘雯
Owner 兰州海关技术中心
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