The Global Analysis Of Gene Expression In Anthurium Responsed To Blight Disease And The Indification Of Resistance-related WRKYs

Bacterial blight, caused by Xanthomonas axonopodis pv.dieffenbachiae (Xad), is the most destructive disease in anthurium. WRKYs transcription factors, have uniquely expanded and evoled in plants to coordinate gene expression. Members of the complex family of WRKY transcription factors play the roles in disease resistance, responses to biotic or abiotic stresses and developmental processes in plants. The global gene expression profiling of Anthurium responsed blight and the identification of WRKYs genes involved in disease resistant, will facilitate revealing the molecular pathology mechanism of Anthurium-pathogen interaction, to improve the resistive breeding. In this study, using Solxea sequencing and bioinformatics technology, the transcriptome of resistant Anthurium’Acropolis’was analyzed and annotated, and used as reference for the digital gene-expression (DGE) analysis about both the foliage and systemic infection. The dynamic expression of genes involed defence pathway srevealed the roles of PTI and ETI immunity system, and hormone-mediated signaling pathways. Moreover, Xad WKRYs were identified. All the results as follow:1.Builded Anthurium cDNA library mix tissues of ’Acropolis’, and77,707Unigenes was obtained by sequencing assembly. Used DGE technologies to study the materials of foliage and systemic infection of Xad, and classifying differential expressed genes, with GO, which suggested that a lot of enrichment-related gene expressed in the immune and stress resistance. In both organizations, PTI related FLS2, FRK1, NDR1and other genes were induced rapidly, and can last for a long time (>12dpi) with a high level of expression; similarly, the pathway components like RPS2, RAR1, HSP90, WRKYs and ETI excited by pathogens-effector were upregulated. Two immune response, PTI and ETI, has coordination on Anthurium disease-resistant and may crosstalk in time and space.2. We analysed comprehensive the of the expression of SA, JA, ET, ABA, GA and other hormone signal-related genes with infection of Anthurium blight. The genes expression of JA biosynthetic accumulation, receptor-binding, and JA downstream-respond genes were up-regulated, indicated the signaling pathway mediated by JA may play an important role in Anthurium disease-resistant; the gene expression that the signaling pathway mediated by SA was down-regulated or no change, indicated the signaling pathway mediated by SA may inactecd in Anthurium disease-resistant because JA and SA may antagoned. In addition, ET signaling pathway may effected systemic resistance by analyzing the gene expression change of the signal-regulated genes and crosstalk gene.The resistance of foliage and systemic down-regulated by ABA needed more experimental evidence.3. In this study, six commonly used candidates including EF1-α, GADPH, UBQ7, His3, TUB and ACTB, were employed to identify the most reliable reference genes. The genes amplification efficiency was assessed by qRT-PCR, and expression stability in the processes of foliage and systemic infection were analyzed on both resistant and suscepti-ble cultivars, then generated dates were evaluated comprehensively using softwares. As a result, the tested genes showed different expressing characters in various samples, among which, the expression of UBQ7and His3was much more stabile in different tissues and varieties, can be recommended as reference gene for data normalization.4. Eight WRKY genes were preliminary identificated with the transcriptome anno-tation information, including20Unigenes and8Contigs, of which12genes cover the full length ORF sequences, and16EST’s sequences. According to the results of the exp-ression profile,we selected10candidate WRKY gene related to disease-resistant.5. The expression analysis of WRKY genes in different tissue like ’Dakoda’ and ’Acropolis’ induced on Xad stress were conducted by qRT-PCR with His3as reference gene. The qRT-PCR results showed that there were four genes U3019, U19502, U12707, CL6222.1of which rapidly improved (1d) relative expression level in Anthurium stems which induced on Xad stress. U13633improved relative expression level in leaves and stems,but no significant changes in ’Dakoda’. Forecasted the relationship between these genes and response of induced on Xad was close. In addition, the analysis of candidate WRKY expression on hormone-induced may initially speculated defense signaling pathways.6. Four genes were cloned the full-length cDNA from material induced on Xad stress by PCR and RACE. The ORF was703bp,1056bp,1266bp,942bp encoding234,351,421,313amino acids,and were validated by amplification of the cDNA and DNA.The results privide a foundation of understanding and the study of the molecular mechanisms of stress anthurium blight.