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Duplex PCR (polymerase chain reaction) primer and method for identifying tomato chlorosis viruses and tomato yellow leaf curl viruses

A technology for tomato yellowing leaf curl and tomato chlorosis virus, which is applied in the field of agricultural biology and achieves the effects of high specificity, improved detection efficiency, and reduced operation steps and reagent consumption.

Inactive Publication Date: 2015-11-25
QINGDAO AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, there is no report on the rapid identification of ToCV and TYLCV using double PCR technology

Method used

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  • Duplex PCR (polymerase chain reaction) primer and method for identifying tomato chlorosis viruses and tomato yellow leaf curl viruses
  • Duplex PCR (polymerase chain reaction) primer and method for identifying tomato chlorosis viruses and tomato yellow leaf curl viruses
  • Duplex PCR (polymerase chain reaction) primer and method for identifying tomato chlorosis viruses and tomato yellow leaf curl viruses

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] The suspected virus plants and healthy plants described in Example 1 are long-term raised plants in the Invasive Biology Laboratory of Qingdao Agricultural University; the primers and methods designed by the present invention are used to detect the viruses on the above plants, and the plants in the greenhouse are ToCV and TYLCV Compound infected plants.

Embodiment 2

[0034] The suspected virus plants described in Example 2 were collected in Shangma District, Qingdao City, Shandong Province in 2014; the above-mentioned virus suspected plants were detected using existing technologies, and some of the plants collected in Shangma were ToCV and TYLCV co-infected plants, and some were TYLCV infected plants alone, but ToCV alone infected plants were not detected.

Embodiment 3

[0035] The suspected virus plants described in Example 3 were collected in Laiyang, Yantai City, Shandong Province in 2014; the above-mentioned virus suspected plants were detected using existing technologies, and some of the plants collected in Laiyang were ToCV and TYLCV co-infected plants, and some were TYLCV infected plants alone, but ToCV alone infected plants were not detected.

[0036] The RNA extraction reagent Trizol described in the examples was purchased from Invitrogen Company, the cDNA synthesis reagent PrimeScriptRTreagentKit was purchased from TaKaRa Company, and other reagents were common commercially available reagents.

[0037] Example 1

[0038] A method for discriminating between tomato chlorosis virus and tomato yellow leaf curl virus, the steps are as follows:

[0039] (1) extract the RNA of ToCV and TYLCV infection virus plant sample, make RNA solution;

[0040] RNA was extracted according to the steps of Trizol (Invitrogen, Carlsbad, CA, USA) reagent, a...

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Abstract

The invention relates to a duplex PCR (polymerase chain reaction) primer and a method for identifying tomato chlorosis viruses and tomato yellow leaf curl viruses. The method comprises the following steps that (1) plant total RNA (ribose nucleic acid) is extracted; (2) the plant total RNA is used as a template for synthesizing cDNA (complementary deoxyribonucleic acid) through reverse transcription; (3) the cDNA is used as a template for performing PCR amplification on a nucleotide sequence coded by ToCV and TYLCV capsid protein; (4) a PCR product prepared in the step (3) is subjected to agarose gel electrophoresis analysis. The invention relates to a PCR primer combination. A duplex PCR method is used for performing ToCV and TYLCV identification; the result is accurate; the method is fast, simple and convenient; the detection cost is reduced; the virus detection efficiency is greatly improved; good application prospects are realized.

Description

technical field [0001] The invention relates to a double PCR primer and a method for distinguishing tomato chlorosis virus and tomato yellow leaf curl virus, belonging to the field of agricultural biotechnology. Background technique [0002] Tomato yellow leaf curl virus (Tomatoyellowleafcurlvirus, TYLCV) and tomato chlorosis virus (Tomatochlorosisvirus, ToCV) are two important viruses on tomato, which pose a serious threat to tomato production. TYLCV is a DNA virus belonging to the genus Begomovirus of the family Geminiviridae. Infected tomato plants are dwarfed, grow slowly or stagnate, and the top leaves are often slightly green, yellow, and smaller. Thickening and hardening of leaves are the primary diseases of tomato in protected areas. ToCV is an RNA virus belonging to the genus Crinivirus of the Closteroviridae family. Infected tomato leaves become chlorotic and yellow, and the veins are dark green. The affected leaves become brittle and easy to break, and the yellow...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/11C12Q1/68
Inventor 李洁孙珊褚栋丁天波
Owner QINGDAO AGRI UNIV
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