CN-122011143-A - Huang-Huai sacculus mildew effector protein EPI3, coding gene and application thereof

Abstract

The invention belongs to the technical field of genetic engineering, and discloses a yellow-Huai sacculus mildew effector protein EPI3 protein, and a coding and application thereof, wherein the amino acid sequence of Huang Huaiqiu sacculus mildew effector protein EPI3 is shown as SEQ ID NO.2, and the nucleotide sequence for coding the gene is shown as SEQ ID NO. 1. The cloning of the sacculus flavescens EPI3 protein gene and the transient expression of tobacco prove that the sacculus flavescens EPI3 protein plays an important role in the process of improving plant disease resistance.

Inventors

• CHEN JIAJIA
• PENG JINFENG
• YU JIAN
• SUN YAYA
• FU DANYANG
• WU XIANGQIAN
• HUANG XIAOXIAO
• ZHENG XIANGRONG
• LI SONG

Assignees

• 江苏农林职业技术学院

Dates

Publication Date

20260512

Application Date

20260205

Claims (10)

1. 1. The yellow-Huai sacculus mildew effector protein EPI3 is characterized in that the amino acid sequence of the Huang Huaiqiu sacculus mildew effector protein EPI3 is shown as SEQ ID No. 2.
2. 2. A nucleic acid or gene encoding the Huang Huaiqiu cyst mould effector protein EPI3 of claim 1, wherein the nucleotide sequence of said nucleic acid or gene is shown in SEQ ID No. 1.
3. 3. An expression cassette, recombinant expression vector, transgenic cell line or transgenic recombinant bacterium comprising a nucleic acid or gene of the Huang Huaiqiu sporangium effector protein EPI3 of claim 2.
4. 4. The method for constructing a recombinant expression vector according to claim 3, wherein the expression vector is obtained by introducing a gene encoding Huang Huaiqiu cyst mold effector protein EPI3 into a plant expression vector pCAMBIA 1300-FLAG.
5. 5. The construction method according to claim 4, wherein the nucleotide sequence of the primer used in the construction process of the expression vector is shown in SEQ ID NO. 3-4.
6. 6. The recombinant bacterium according to claim 3, wherein the recombinant expression vector according to claim 3 is inserted into E.coli and the recombinant bacterium is obtained by screening.
7. 7. The transgenic recombinant strain of claim 6, wherein the escherichia coli is a DH5 a competent cell.
8. 8. Use of Huang Huaiqiu a cyst mold effector protein EPI3 of claim 1, a nucleic acid or gene of claim 2, an expression cassette, a recombinant expression vector, a transgenic cell line, or a transgenic recombinant bacterium of claim 3 in the preparation of a reagent for inducing plant allergic reactions.
9. 9. Use of Huang Huaiqiu a cyst mold effector protein EPI3 of claim 1, a nucleic acid or gene of claim 2, an expression cassette, a recombinant expression vector, a transgenic cell line, or a transgenic recombinant bacterium of claim 3, in the preparation of a reagent for increasing plant resistance.
10. 10. The use according to claim 9, wherein said increasing plant resistance is increasing the resistance of a plant to phytophthora capsici infection.

Description

Huang-Huai sacculus mildew effector protein EPI3, coding gene and application thereof Technical Field The invention belongs to the technical field of genetic engineering, and particularly relates to a balloon mold effect protein EPI3 of Huang Huai, and a coding gene and application thereof. Background Huang Huaiqiu sporocyst mould (Globisporangium huanghuaiense), the synonym is Pythium ultimum (Pythium huanghuaiense), is an earth-borne pathogenic bacterium, can cause root rot and damping-off of various crops such as wheat, rice, soybean and the like, causes the reduction of the emergence rate of the crops, the resistance of plant growth and even death, and causes serious economic loss to agricultural production. During the infection process, part of plant pathogenic bacteria can secrete a plurality of effector proteins into host tissues, and the proteins are key factors for regulating the plant immune response and mainly can be divided into two types, namely apoplast effector proteins (acting on the extracellular space of a plant) and cytoplasmic effector proteins (entering the plant cell to play a role) according to the action sites. Depending on the function, effector proteins can be divided into elicitors (which induce plant immune responses) and suppressors (which suppress plant immunity), which play opposite roles in pathogen-host interactions. Wherein the exciton is recognized by a pattern recognition receptor on the surface of the plant cell, thereby activating "pattern-triggered immunity" (PTI). The exciton has the characteristics of inducing plant disease resistance, being environment-friendly and the like, and has become an important direction for developing traditional chemical pesticide substitutes. EPI effector protein (Extracellular Protease Inhibitor, EPI) is a pathogenic effector molecule identified from oomycete secretion groups such as phytophthora infestans reported by previous researches, the nitrogen end of a protein sequence codes a signal peptide, and the protein sequence is formed by serially connecting Kazal type structural domains with protease inhibition activity, is widely existing in phytophthora and has a conserved function, and related functional reports are not found in sacculus mould at present. Analyzing immune activation mechanism of EPI effect protein, developing recombinant EPI protein capable of triggering plant allergic reaction (HR) with high efficiency, and inducing plants to generate broad-spectrum disease resistance by activating HR to rapidly start plant immune response, thus having great theoretical value and application prospect for developing environment-friendly oomycete disease prevention and control strategy. Disclosure of Invention Object of the invention A first object of the present invention is to provide a Huang Huaiqiu cyst EPI effector protein that can induce plant allergic reactions (HYPERSENSITIVE RESPONSE, HR). A second object of the present invention is to provide a nucleic acid or gene encoding the above Huang Huaiqiu cyst membrane EPI effector protein. The third object of the present invention is to provide an expression cassette, a recombinant expression vector, a transgenic cell line or a transgenic recombinant bacterium containing the above nucleic acid or gene, and construction methods of the recombinant expression vector and the transgenic recombinant bacterium. The fourth object of the invention is to provide the use of the Huang Huaiqiu sporangium EPI effector protein, nucleic acid or gene, expression cassette, recombinant expression vector, transgenic cell line or transgenic recombinant bacterium. In order to achieve the above purpose, the present invention adopts the following technical scheme: the amino acid sequence of the Huang Huaiqiu saccorium vulgare effector protein EPI3 is shown as SEQ ID No. 2. The nucleotide sequence of the nucleic acid or the gene for encoding Huang Huaiqiu cyst effector protein EPI3 is shown as SEQ ID NO. 1. The invention relates to an expression cassette, a recombinant expression vector, a transgenic cell line or a transgenic recombinant bacterium containing the nucleic acid or the gene of Huang Huaiqiu cyst membrane effector protein EPI 3. The invention relates to a construction method of a recombinant expression vector, which is obtained by introducing a gene for encoding Huang Huaiqiu cyst membrane EPI effector protein into a plant expression vector pCAMBIA 1300-FLAG. The nucleotide sequence of the primer used in the construction process of the expression vector is shown in SEQ ID NO. 3-4. The transgenic recombinant strain is obtained by inserting the recombinant expression vector into escherichia coli and screening. Wherein the E.coli is DH5 alpha competent cells. The Huang Huaiqiu cyst effector protein EPI3, nucleic acid or gene, expression cassette, recombinant expression vector, transgenic cell line or transgenic recombinant bacteria are applied to the preparation of plant allergic reaction inducing