Preparation method and application of antiviral potato plant

A potato and anti-virus technology, which is applied in the field of plant anti-virus genetic engineering, can solve the problem of less gene editing in potatoes, and achieve the effect of reducing losses and mitigating injuries

Pending Publication Date: 2022-08-05
SHANDONG AGRICULTURAL UNIVERSITY
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  • Abstract
  • Description
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Problems solved by technology

[0007] At present, there are few

Method used

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  • Preparation method and application of antiviral potato plant
  • Preparation method and application of antiviral potato plant
  • Preparation method and application of antiviral potato plant

Examples

Experimental program
Comparison scheme
Effect test

Embodiment

[0054] 1. Design of target gRNA

[0055] Using the CRISPR design software, the gRNA target was designed with the StPCaP1 gene of the NCBI database, and 7 guideRNA (gRNA) sequences were obtained (as shown in Table 1). The GenBank accession number of StPCaP1 mRNA is XM_006352559.2, and the nucleotide sequence of StPCaP1 gene is shown in Seq ID No.18.

[0056] The design principles are:

[0057] (1) The length of the target site sequence is 20bp;

[0058] (2) The target site sequence is followed by three bases of NGG (N is any base);

[0059] (3) It is best to design the target site at the front of the gene exon.

[0060] Table 1 guide RNA sequences

[0061]

[0062] 2. Construction of CRISPR / Cas9 expression vector

[0063] 1. Primer Design

[0064] The above sequences numbered 1 and 2, namely gRNA1 and gRNA2, were constructed on a knockout vector, named StPCaP-g12 (primers 1, 2);

[0065] The above-mentioned sequences numbered 3 and 4, namely gRNA3 and gRNA4, were cons...

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Abstract

The invention belongs to the field of plant antiviral gene engineering, and discloses a preparation method and application of an antiviral potato plant. The preparation method comprises the following steps: (1) constructing a carrier by taking pH SE401 as a skeleton, and introducing insertion, deletion or substitution mutation into nucleotides from the 762nd site to the 781th site of an StPCaP1 gene; (2) transforming the obtained vector into agrobacterium, transforming the screened agrobacterium positive transformant into potatoes by using an agrobacterium-mediated leaf disc transformation method, and screening potato plants with StPCaP1 gene knockout; and (3) performing friction inoculation on the potato virus until the potato plant with the StPCaP1 gene knocked out, and detecting the anti-virus capability of the potato plant to obtain the anti-virus potato plant. The anti-virus potato plant prepared by the preparation method can obviously inhibit the infection of potato viruses, especially potato Y viruses, obviously reduce the damage of the plants infected by the potato Y viruses, and obviously reduce the loss.

Description

technical field [0001] The invention belongs to the field of plant antiviral genetic engineering, and in particular relates to a preparation method and application of antiviral potato plants. Background technique [0002] Virus disease is an important disease on crops, causing huge losses to agricultural production. Due to the variety of crop virus diseases and the complicated transmission routes, there are no varieties with immunity or high resistance to virus diseases in production, and there are no drugs with specific effects on virus diseases on the market, so the prevention and control of virus diseases is very difficult. [0003] There are many kinds of plant viruses that cause viral diseases, such as tobacco mosaic virus, cucumber mosaic virus, potato virus and so on. Potyvirus genus (Potyvirus) has a wide host range and can infect a variety of Solanaceae crops including potato, tobacco, tomato and pepper, causing huge economic losses. [0004] Potato is the fourth ...

Claims

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Application Information

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IPC IPC(8): C12N15/84C12N15/29C12N15/55C12N15/113C12N1/21A01H5/00A01H6/82
CPCC12N15/8218C12N15/8283C07K14/415C12N9/22
Inventor 耿超李向东房乐田延平
Owner SHANDONG AGRICULTURAL UNIVERSITY
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