Phytophthora parasitica avirulence gene PnAvr8, encoding protein and application thereof

A technology of Phytophthora parasiticus and avirulent gene, applied in the field of agricultural biology, can solve the problem of no avirulent gene, and achieve the effects of avoiding breeding redundancy, assisting the cloning of disease resistance genes, and stable results

Active Publication Date: 2022-07-29
TOBACCO RES INST CHIN AGRI SCI ACAD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, for Phytophthora parasiti...

Method used

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  • Phytophthora parasitica avirulence gene PnAvr8, encoding protein and application thereof
  • Phytophthora parasitica avirulence gene PnAvr8, encoding protein and application thereof
  • Phytophthora parasitica avirulence gene PnAvr8, encoding protein and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] like figure 1 As shown, the tested tobacco varieties Beinhart1000 (BH) and Xiaojinjin 1025 (XHJ) were provided by the germplasm resource bank of the Institute of Tobacco Research, Chinese Academy of Agricultural Sciences. After years of experimental investigation and observation of BH, both new plants and perennials showed high resistance, and XHJ was identified as a high-susceptibility variety. The pathogen of black shank tested was Phytophthora parasiticus No. 0 physiological race, which was preserved by the Tobacco Research Institute of the Chinese Academy of Agricultural Sciences. When the hydroponic tobacco seedlings grow to the stage of 4 true leaves, place BH and XHJ in a petri dish covered with Phytophthora parasitica hyphae, and take samples before inoculation and 6, 12, 24, and 60h after inoculation, and take a total of 5 times. , for transcriptome analysis.

[0030] According to the gene expression of Phytophthora parasiticus at different time points in the...

Embodiment 2

[0032] Construction of transient expression vector: 17 candidate RXLR effectors in Example 1 were synthesized, and 17 RXLR effectors were constructed into the expression vector PVX virus plant expression vector pGR106 by the Gateway method. The sequencing was correct, and the positive clones were confirmed and transferred into Agrobacterium GV3101 save in.

[0033] Toothpick puncture screening: Agrobacterium containing the RXLR effector gene was activated on LB plate medium and cultured at 28°C for 48h. Pick a single clone and put it into 1mL LB liquid medium, culture it overnight at 28°C, 200rpm, and culture it to OD. 600 The value is about 1.0. 500 μL of bacterial liquid was spread on LB solid medium, cultured at 28°C overnight, and a small amount of Agrobacterium was used to puncture tobacco leaves with a sterile toothpick. pGR106-GFP was the negative control and pGR106-CRN2 was the positive control. Each plant had 3 leaves, and each leaf had 3 puncture points, which wer...

Embodiment 3

[0037] BlastP searches were performed on various oomycete genomic data through the National Center for Biotechnology Information (NCBI) to determine whether PnAvr8 homologous genes are present in other organisms. It was found that if image 3 As shown, PnAvr8 has highly similar homologous proteins in different P. Very high interspecies specificity. The secondary structure of PnAvr8 was predicted through the protein structure prediction website and the amino acid sequence was analyzed. It was found that there were two α-helix structures downstream of the signal peptide of PnAvr8.

[0038]

[0039]

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Abstract

The invention belongs to the technical field of agricultural biology, and particularly relates to a phytophthora parasitica avirulence gene PnAvr8, an encoding protein and application of the phytophthora parasitica avirulence gene. On the basis of whole genome sequencing of a phytophthora parasitica strain ASM148298v1, 17 RXLR effect factors which are remarkably up-regulated in the early infection stage are preliminarily predicted through transcriptome data of roots of disease-resistant and susceptible tobacco varieties infected by phytophthora parasitica. 17 RXLR effect factors are constructed into a PVX virus plant expression vector pGR106, and one effect factor PnAvr8 is screened from tobacco germplasm resources through a toothpick puncture method, can excite the HR reaction in a nicotiana aureoides resistant material, and cannot excite the HR reaction in common nicotiana aureoides disease-resistant and susceptible materials. The gene can be used as a resistance screening tool for breeding of tobacco resistant to phytophthora parasitica, and tobacco resources with corresponding anti-disease genes can be rapidly determined. In the process of tobacco crossbreeding and population construction, the PnAvr8 can also be used as an identification tool, so that the tobacco resistance breeding population construction is more convenient.

Description

technical field [0001] The invention belongs to the technical field of agricultural biotechnology, and specifically relates to a Phytophthora parasiticus avirulence gene PnAvr8, an encoded protein and applications thereof. Background technique [0002] The genus Phytophthora contains many serious pathogens that can infect all dicotyledonous plants and some monocotyledonous plants. Among them, tobacco black shank disease caused by Phytophthora nicotianae is one of the most devastating diseases in tobacco production, and it is highly pathogenic to many Solanaceae crops, for example, it can cause potato tubers to rot, sweet peppers , Lycium barbarum root rot and tomato brown rot. It ranks eighth in the latest Oomycetes hazard ranking, and due to its higher optimum growth temperature, its host range still has a tendency to expand significantly with global warming. [0003] Phytophthora pathogens secrete effectors, which create a suitable environment for the invasion and prolif...

Claims

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Application Information

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IPC IPC(8): C07K14/37C12N15/31C12N15/84C12Q1/18C12R1/645
CPCC07K14/37C12N15/8205C12Q1/18Y02A50/30
Inventor 孟鹤程立锐杨爱国王元英王红万智焱姜自鹏
Owner TOBACCO RES INST CHIN AGRI SCI ACAD
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