Identification Of A Defence Gene PR4A From Citron C-05 In Response To Xanthomonas Citri Subsp.citri Infection

As a worldwide quarantine disease,citrus canker can bring great damages to all Commercial citrus genotypes.Citrus canker is one of the most devastating disease that it can cause citrus leaves to fall off early.To date,with no cure for citrus canker,the spread of citrus canker can only be controlled by using copper spray to inhibit the growth of Xcc.Thus,disease resistance breeding has become the key measure to help the citrus industry resist citrus canker.With hundreds of citrus genotypes were screened worldwide,there was no resistance has been obtained,despite the field tolerance described for some citrus genotypes.Citron C-05 with stable resistance to citrus canker has been recognized in recent ten years at our lab.The mechanism of resistance in Citron C-05 to citrus canker must be proved to apply Citron C-05to disease resistance breeding.In this paper,to screen the main PR4 gene for the resistance of Citron C-05 to citrus canker and to analyze the function of PR4 gene in the resistance of Citron C-05 to citrus canker,the response to Xcc in Citron C-05 was analyzed with the control of the response to Xcc in susceptible citrus genotypes.The main results were as follows:1.The transcriptome data showed PR1-1?PR2-1?PR2-3?PR3-1 and PR8-2 gene may participate in resistance of Citron C-05 to Xcc.And PR4-5 was specifically up-regulated in Citron C-05 leaves infected with Xcc,and it keep the high expression until 12 dpi.The result suggested that PR4A may be a key gene involved in the resistance of Citron C-05 to citrus canker.2.There are 5 tandem PR4 gene on chromosome 8 of citrus sinensis were found.The PR4-3 and PR4-4 are both Class?PR4 proteins with Hevein domains,while PR4-1?PR4-2and PR4A are all Class?PR4 proteins without Hevein domains;PR4A gene had significantly upregulated in transcriptome of Citron C-05 leaves which injected with Xcc.To analyze the responses of PR4 genes to Xcc,Xcc was inoculated to leaves of citron C-05 and susceptible citrus genotypes.The result suggested that the expression of PR4A gene in Citron C-05 was significantly higher than that in susceptible genotypes at the same time.That showed PR4A gene was involved in the resistance of Citron C-05 to Xcc.3.After the alignment of genes sequence,there were 2 exons and 1 intron had been found in PR4As of Citron C-05 and susceptible genotypes.The 2 exons had the same among all genotypes,while the intron differences caught the difference on length of genes.Alignment of the amino acid sequences of PR4As found that the Citron C-05 PR4A protein contained 2specific amino acid sites(95^th:T/A;97^th:E/Q)located in Barwin domain which may cause the functional differences of PR4As protein between Citron C-05 and susceptible genotypes.However,the secondary structure and tertiary structure of PR4A proteins showed no difference.4.We selected CsPR4A and CmPR4A to further evaluate their capacity to enhance bacterial resistance using Agrobacterium-mediated transient expression assays.Both CsPR4A and CmPR4A,the two protein reduced the growth of Xcc suggested that the both proteins had the resistance to citrus canker.But we can not sure if there were differences between the function of CsPR4A and CmPR4A because of the higher expression of PR4A in CmPR4A transient expression leaves.