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Primer, kit and identification method for potato X virus identification

An identification method and potato technology, applied in the biological field, can solve the problems of yield reduction, heavy mosaic leaves, dwarf plants, etc.

Inactive Publication Date: 2017-11-17
陈定虎
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the field, PVX infects potato plants, causing light mosaic or latent symptoms. If infected for many years, it will easily cause heavy mosaic or plant dwarfing, resulting in 10%-80% yield reduction

Method used

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  • Primer, kit and identification method for potato X virus identification
  • Primer, kit and identification method for potato X virus identification
  • Primer, kit and identification method for potato X virus identification

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0077] A kit for identifying potato virus X RT-LAMP, the kit includes:

[0078]

[0079] Among them, the primer sequence:

[0080] PVX-F3:5'-ACAGGCTGCTTGGGACTT-3'

[0081] PVX-B3:5'-GTTAGCAGGTGGACTGTTGT-3'

[0082] PVX-FIP:5'-ATGCCGTTGGAATAGGGACCTG-GCTGATGTAGGATCATCCGC-3'

[0083] PVX-BIP: 5'-AGCAGAGCTAGACTGGCAGCA-CCATACCACTGGGGCATAC-3'.

Embodiment 2

[0085] A kit for identifying potato virus X RT-LAMP, the kit includes:

[0086]

[0087] Among them, the primer sequence:

[0088] PVX-F3:5'-ACAGGCTGCTTGGGACTT-3'

[0089] PVX-B3:5'-GTTAGCAGGTGGACTGTTGT-3'

[0090] PVX-FIP:5'-ATGCCGTTGGAATAGGGACCTG-GCTGATGTAGGATCATCCGC-3'

[0091] PVX-BIP: 5'-AGCAGAGCTAGACTGGCAGCA-CCATACCACTGGGGCATAC-3'.

[0092] The 2× reaction solution buffer described in the present invention is made up of following components:

[0093]

[0094]

[0095] The enzyme solution is a mixture of Bst DNA polymerase and AMV reverse transcriptase.

[0096] The fluorescent visual detection reagent contains calcein fluorescent dye.

Embodiment 3

[0098] A kind of potato virus X identification method, comprises the following steps:

[0099] A. Plant tissue total DNA extraction;

[0100] B. LAMP detection:

[0101] The kit described in Example 2 was used to configure the pre-reaction solution, and 5 μL of DNA of the sample to be tested was added to the pre-reaction solution to make the total amount reach 25 μL; the control group added 5 μL of deionized water to the pre-reaction solution; positive For the control, add 5 μL of virus-carrying DNA to the pre-reaction solution; use a pipette to mix by suction and drainage, or close the lid and tap lightly to make the solution fully mixed and then centrifuge instantaneously, avoiding bubbles during mixing; place the mixture in the reaction well, React at a constant temperature of 65°C for 90 minutes, and finally keep it at 80°C for 5 minutes to end the reaction;

[0102] C. LAMP result judgment

[0103] After the amplification reaction is over, use an ultraviolet irradiatio...

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Abstract

The invention discloses a primer, a kit and an identification method for the identification of a potato X virus (RT-LAMP). The kit comprises two reaction buffer solutions, a primer (FIP), a primer (BIP), a primer (F3), a primer (B3), an enzyme solution, a fluorescence visual detection reagent, and deionized water. The provided primer, kit and identification method can overcome the shortages of the prior art, and have the advantages of rapidness, simpleness, convenience, efficiency, high accuracy, strong specificity, and high sensitivity.

Description

technical field [0001] The invention relates to a primer, a kit and a detection method for identifying potato virus X RT-LAMP, belonging to the field of biotechnology. Background technique [0002] Potato virus X, PVX, also known as Potato latent virus or Potato mild mosaic virus, is a model member of Potatovirus X. It is a linear virus composed of a positive strand RNA. About 6.4kbp, the virion is in the shape of a long rod, length×width=515nm×13nm. Under natural conditions, mechanical transmission mainly depends on different parts of the host plant and contact friction between the host plants. The symptoms are mild when PVX is infected alone, and the symptoms are aggravated when combined with Potatovirus Y, PVY, Tobacco veinmottling virus, TVMV, Tobacco etch virus, TEV and other Potatovirus Y viruses , causing serious economic losses. [0003] PVX occurred in all potato Solanum tuberosum production areas, but there were great differences in the infection of different cu...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/68C12N15/11C12N15/10C12R1/94
CPCC12Q1/6806C12Q1/6844C12Q1/701C12Q2531/119C12Q2563/107C12Q2563/143C12Q2563/149
Inventor 刘洪义陈定虎陈文聪刘运张静苏斐
Owner 陈定虎
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