Study On Resistance Mechanism In HRF1Transgenic Rice To Xoo And Function Analysis Of Rice Resistance-Related Gene OSCYP71Z2

Bacterial blight is one of the most destructive rice diseases, which caused by Xanthomonas oryzae pv. oryzae (Xoo), and results in10%to80%yield losses, endangering worldwide food security. Presently heavily application of chemical pesticides for controling disease has seriously damaged ecological environment. Meanwhile, easy lost of rice variety resistance to disease posed higher demond for rice breeding. An economically effective and environmentally sound approach to control disease is the utilization of cultivars that possessed durable, broad-spectrum resistance to disease by biotechnology. Identification of rice resistance-related genes is the base to cultivate the new rice disease-resistant varieties. The exploration of gene code elicitor protein with potential application value provides available resource for controlling disease by inducing multiple defense responses.Harpinxoo proteins from the Xoo stain JXOIII increase plant disease resistance by activating multiple defense responses in plants, such as systemic acquired resistance and hypersensitive response. However, it is unclear whether phytoalexin production and ROS burst are involved in the disease resistance conferred by the expression of the harpinxoo protein in rice. In preliminary studies, our group showed some hrfl (code harpinxoo protein)-transformed rice T3homozygote increased resistance to Xoo. On the basis, in this article hrfl-transformed rice T3homozygote NJH12has been choiced for analyzing the resistance mechanism to Xoo. In this article, ectopic expression of hrfl in rice enhanced resistance to bacterial blight. Accompanying the activation of genes related to the phytoalexin biosynthesis pathway in hrfl-transformed rice, phytoalexins quickly and consistently accumulated concurrent with the limitation of bacterial growth rate. Moreover, the hrfl-transformed rice showed an increased ability for ROS scavenging and decreased hydrogen peroxide (H2O2) concentration. Furthermore, the localization and relative quantification of silicon deposition in rice leaves was detected by scanning electron microscopy (SEM) and energy-dispersive X-ray spectrometer (EDS). Finally, the transcript levels of defense response genes increased in transformed rice. These data are significant because they provide evidence for a better understanding the role of defense responses in the incompatible interaction between Xoo and hrf1-transformed plants. The data also supply an opportunity for generating nonspecific resistance to pathogens.Based on the transcriptome profile of the transgenic rice line NJH12expressing hrfl from Xoo, a resistance-related gene showing114.6-fold increase of expression level was identified. The gene is the cytochrome P450gene, named as Oscyp71Z2analyzed by bioinformation, which belongs to the CYP71Z subfamily. Oscyp71Z2CDS, Oscyp71Z2RNAi target fragment and promoter sequence have cloned from rice cultivar Nipponbare transcript and genome, respectively make transformation vectors for overexpression, RNAi and Oscyp71Z2expression pattern, then transformed into rice Nipponbare genome mediated by Agrobacterium tumefaciens. The results from normal PCR and Southern blot showed T-DNA has inserted into rice genome. The data from Northern blot and Western blot showed the transcript and protein level in Oscyp71Z2-overexpressiong plant was higher than that in wild type Nipponbare. On the contrary, the protein level in Oscyp71Z-RNAi plant was lower than that in wild type, the tanscript hasa no detected in both. A strong GUS signal was detected mainly in the leaves, nodes, primary roots and glume. Overexpression of Oscyp71Z2enhanced resistance to Xoo at the booting stage, and inhibited multiplication of Xoo PXO99A. The accumulation of phytoalexins was rapidly and strongly induced in Oscyp71Z2-overexpressing plants, and the transcript levels of genes related to the phytoalexin biosynthesis pathway were elevated. The H2O2concentration in Oscyp71Z2-overexpressing plants was reduced in accordance with the increase in ROS-scavenging ability due to the induction of superoxide dismutase (SOD) as well as peroxidase (POD) and catalase (CAT) activation. We also showed that suppression of Oscyp71Z2had no significantly effect on disease resistance to Xoo in rice, although inhibited part phytoalexin production. These results demonstrated that Oscyp71Z2plays an important role for bacterial blight resistance by regulating the diterpenoid phytoalexin biosynthesis and H2O2generation.