阅读说明：本技术 野生柑橘资源AbTFIIAγ基因及其在柑橘抗溃疡病中的应用 (Wild citrus resource AbTFIIA gamma gene and application thereof in citrus canker resistance ) 是由 徐强 汤小美 于 2020-04-10 设计创作，主要内容包括：本发明属于植物基因工程领域,其公开了一种野生柑橘资源AbTFIIAγ基因及其在柑橘抗溃疡病中的应用。本发明从野生资源酒饼簕(Atalantia buxifolia,柑橘的野生近缘种)中克隆到TFIIAγ基因,其核苷酸序列如SEQ ID NO：1所示,其编码的蛋白序列如SEQ ID NO：3所示,不能被溃疡病菌携带的效应子TALEs结合和利用,进一步明确了影响TAL效应子与TFIIAγ互作的重要氨基酸突变位点。本发明为柑橘溃疡病抗性品种培育提供了重要基因资源和靶位点。(The invention belongs to the field of plant genetic engineering, and discloses a wild citrus resource AbTFIIA gamma gene and application thereof in citrus canker resistance.A nucleotide sequence of the gene is shown as SEQ ID NO: 1, and a coded protein sequence of the gene is shown as SEQ ID NO: 3, so that an effector TA L Es which cannot be carried by canker pathogenic bacteria is combined and utilized, and important amino acid mutation sites influencing interaction of the TA L effector and the TFIIA gamma are further defined.)

1. A wild citrus resource AbTFIIA gamma gene has a nucleotide sequence shown as SEQ ID NO: 1 is shown.

2. A nucleotide sequence of the coding region of the gene of claim 1, which sequence is as set forth in SEQ ID NO: 2, respectively.

3. An AbTFIIA gamma protein sequence encoded by the gene of claim 1, wherein the amino acid sequence is as shown in SEQ ID NO: 3, respectively.

4. Use of the wild citrus resource AbTFIIA γ gene of claim 1 in citrus anti-ulcer breeding.

Technical Field

The invention relates to the field of plant genetic engineering, in particular to a wild citrus resource AbTFIIA gamma gene and application thereof in resisting canker of citrus.

Background

Canker is one of the citrus devastating diseases, and seriously threatens the development of the citrus industry. Most citrus cultivars are susceptible to canker, and necrotic spots can be formed in fruits and leaves of the citrus cultivars after the citrus cultivars get ill, so that the dropping of the fruits and the leaves is accelerated, and the fruit yield and the commodity value are seriously influenced. The prevention and treatment of the canker in the production mainly uses bactericide, which not only increases the planting cost; moreover, the citrus is used as perennial crops, and the use of chemical agents for one year brings risks to the safety and green ecology of products. At present, no commercial citrus varieties which can completely resist canker are found at home and abroad. Therefore, the development of resistance genes and their application in resistance breeding have become an urgent research for the citrus industry.

The Xanthomonas is used for releasing a pathogen transcription activation effector (TA L Es) to plant cells through a three-type secretion system and finally entering cell nuclei, the TA L Es is combined with a promoter of a disease-sensitive gene to activate the expression of the disease-sensitive gene, and the TA L E structure contains a transcription factor binding structural domain (TFB) which is very important for the pathogenicity of the Xanthomonas.

China is one of the origins of citrus, has abundant wild resources, and provides a material basis for excavating resistance gene resources. The panel evaluated the resistance to canker disease based on the collection of wild resources and identified a source of anti-canker disease. And natural variation of TFIIA gamma is searched in the wild resources by applying a genomics tool, and resistance genes are excavated and identified, so that gene resources are provided for culturing citrus canker resistant varieties.

Disclosure of Invention

The invention aims atThe invention provides a wild citrus resource AbTFIIA gamma gene and application thereof in citrus canker resistance, wherein the invention discovers that Acanthopanax trifoliatus (Atalantia buxifolia) has stronger resistance to citrus canker, clones the Acanthopanax trifoliatus TFIIA gamma gene (named as AbTFIIA gamma), discovers that transcription factor binding motifs (TFBi and TFBii, which are abbreviated as TFBs) on TA L effector of 2 citrus canker germs can not be combined with AbTFIIA gamma gene, and on the contrary, TFBs can stably interact with TFIIA gamma gene (named as CsIIA gamma) of a control susceptible variety sweet orange^L81VInstead, after mutating AbTFIIA gamma (81V, 87I, 90N) to the (81L, 87L, 90T) amino acid corresponding to CsTFIIA gamma, the mutation at amino acid position 81 results in pathogenic bacteria TFBs being able to bind to AbTFIIA gamma^V81LCombining; whereas the amino acid mutations at positions 87 and 90 did not have such changes. The research result reveals that AbTFIIA gamma can be used as a candidate gene for improving the citrus canker resistant variety, and the 81 th amino acid coded by the gene is a key site for the interaction between pathogenic bacteria and host plants.

In order to achieve the purpose, the invention relates to a wild citrus resource AbTFIIA gamma gene, the nucleotide sequence of which is shown as SEQ ID NO: 1 is shown.

The nucleotide sequence of the coding region of the wild citrus resource AbTFIIA gamma gene is shown as SEQ ID NO: 2, respectively.

The amino acid sequence of the AbTFIIA gamma amino acid coded by the wild citrus resource AbTFIIA gamma gene is shown as SEQ ID NO: 3, respectively.

The invention also provides application of the wild citrus resource AbTFIIA gamma gene in breeding of citrus canker resistance.

The invention has the beneficial effects that:

the invention discloses an application of orange wild resource Acanthopanax trifoliatus AbTFIIA gamma gene in orange resistanceThe invention finds that citrus susceptibility gene CsTFIIA gamma stably interfering with the interaction of the TFBs structural domain of ulcer pathogenic bacteria increases the resistance of the citrus susceptibility gene CsTFIIA gamma to ulcer, simultaneously finds that the TFBs structural domain can not be combined with AbTFIIA gamma, and when the 81 st leucine (L) of susceptibility allele CsTFIIA gamma is replaced by the 81 st valine (V) of disease-resistant allele AbTFIIA gamma, the TFBs and the CsIIA TFgamma are combined^L81VThe binding capacity of (A) is weakened, and the key role of the variation of the 81 th amino acid on the interaction of pathogenic bacteria and plant hosts is determined. The variation information can be applied to the improvement of the citrus canker-resistant varieties.

Drawings

FIG. 1 is a graph showing the results of an evaluation test of resistance to canker by different wild citrus resources;

FIG. 2 is a graph showing the results of an evaluation test of resistance of sweet orange stably interfering with CsTFIIA gamma gene against ulcer disease;

FIG. 3 is a graph showing the results of the interaction between the domains of TFBs and AbTFIIA γ carried by Citrus canker; TFBs are unable to interact with AbTFIIA γ;

FIG. 4 is a graph showing the results of the interaction between TFBs domains carried by Citrus canker pathogen and mutant CsTFIIA γ and AbTFIIA γ; TFBs and mutant CsTFIIA gamma^L81VThe binding capacity is reduced, and the AbTFIIA gamma can be combined with the mutant AbTFIIA gamma^V81LAnd (4) interacting.

Detailed Description

The present invention is described in further detail below with reference to specific examples so as to be understood by those skilled in the art.