Multiple detection method and kit for simultaneously detecting three sweet potato viruses

A detection kit and technology for sweet potato virus, applied in microorganism-based methods, biochemical equipment and methods, recombinant DNA technology, etc., can solve the problem of not being able to detect more than two sweet potato viruses at the same time, saving costs and avoiding crossover. Pollution, the effect of reducing the number of operations

Pending Publication Date: 2022-07-29
HENAN UNIV OF SCI & TECH +1
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Problems solved by technology

[0003] In order to overcome the problem that two or more sweet potato viruses cannot be detected simultaneously in the background technology, the present invention discloses a multiple detection method and a kit for simultaneously detecting three sweet potato viruses

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  • Multiple detection method and kit for simultaneously detecting three sweet potato viruses
  • Multiple detection method and kit for simultaneously detecting three sweet potato viruses
  • Multiple detection method and kit for simultaneously detecting three sweet potato viruses

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Embodiment Construction

[0035] The present invention can be explained in detail through the following examples, and the purpose of disclosing the present invention is to protect all technical improvements within the scope of the present invention.

[0036] combined with Figures 1 to 4 , a multiplex RT-PCR detection kit for simultaneous detection of 3 sweet potato viruses, the kit includes 6 1.5mL centrifuge tubes, respectively equipped with 2.5mM 10× reaction buffer, 1U / μL Taq enzyme 1.5 mL, upstream primers of 3 sweet potato viruses 10 μM, downstream primers of 3 sweet potato viruses 10 μM, pure water 1.5 mL, 15 μg / mL positive standard 0.5 mL; as required, the 10× reaction buffer is Tris-HCl 100 mM , KCl 500mM and dNTPs; Described positive standard substance is to utilize the CP gene full-length primers of described 3 kinds of sweet potato viruses to carry out PCR amplification respectively, the amplification product is sequenced, and the CP gene of the obtained positive sample carries out the cons...

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Abstract

The invention relates to the technical field of sweet potato virus detection, in particular to a multiple detection method and a kit for simultaneously detecting three sweet potato viruses. The kit comprises a 10 * reaction buffer solution, Taq enzyme, upstream primers of the three sweet potato viruses, downstream primers of the three sweet potato viruses and a positive standard substance. The three sweet potato viruses are a sweet potato chlorotic dwarf virus, a sweet potato pinniform mottle virus and a sweet potato latent virus respectively; the detection method comprises the following steps: (1) extracting total RNA (Ribonucleic Acid) of a sample to be detected, and carrying out reverse transcription to obtain cDNA; (2) PCR amplification; (3) taking 5 mu L of the PCR amplification product in the step (2), and performing electrophoresis on 2% agarose gel containing 5 mu g/ml of ethidium olfactory (EB); (4) result judgment: PCR target amplified fragments are respectively 150bp, 280bp and 480bp; according to the invention, three kinds of sweet potato viruses can be simultaneously detected through one PCR reaction, the operation frequency is reduced, cross contamination is avoided, the usage amount of reagents, primers, templates, consumables and the like is also reduced, and time, labor and cost are saved.

Description

technical field [0001] The invention relates to the technical field of sweet potato virus detection, in particular to a multiple detection method and a kit for simultaneously detecting three kinds of sweet potato viruses. Background technique [0002] As is known, sweet potato is a plant that utilizes tuberous roots to carry out asexual reproduction, which is extremely susceptible to virus infection in the cultivation process, and has a great impact on the yield and quality of sweet potato after virus infection; At present, mixed infection of various viruses has been detected in Guangdong, Fujian, Jiangsu, Sichuan, Beijing, Shandong and other places. It is reported that the loss caused by sweet potato virus in China is as high as 4 billion yuan every year. One of the important limiting factors for potato latent virus, SPLV), etc.; and the common situation is the compound infection of multiple sweet potato viruses, which greatly affects the improvement of sweet potato yield ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/686C12N15/11C12R1/94
CPCC12Q1/701C12Q1/686C12Q2600/16C12Q2537/143C12Q2565/125
Inventor 张小梅李广录侯文邦
Owner HENAN UNIV OF SCI & TECH
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