Transcriptional profiling of Cellulase induced hypersensitive reaction like response in rice. Oryza sativa

Cellulase, a Type II secretion system secreted protein of Xanthomonas oryzae pv. oryzae (Xoo; the casual of bacterial leaf blight of rice) is a potent inducer of rice defense responses such as hypersensitive response like reactions (HR), callose depositions, cell death associated with nuclear fragmentations and impart functional resistance against further Xoo inoculation In order to understand the molecular events associated with cellulase induced HR in rice, whole genome transcriptional profiling was performed using Affymetrix Rice GeneChips Keywords: Expression profiling of a hypersensitive reaction like response Overall design: Leaves of 20 days old green house grown susceptible rice cultivar (Taichung Native-1; TN-1) were infiltrated with either 30-40μl of purified Xoo cellulase (5μg/ml) or with buffer (10mM potassium phosphate buffer pH 6.0) alone (as described in Jha et al. 2007; MPMI vol 20, pp 31-40). The plants were shifted to a growth chamber (28oC; 80% relative humidity; 12/12h light/dark cycle) 48 hours before the treatment. 20-30 leaf pieces covering the infiltrated zone from each of the treatments were harvested 12 h after infiltration. Total RNA isolated from the pooled samples was subjected for expression analysis using Affymetrix GeneChip System. The experiment was repeated with three different biological replicates using RNA isolated from three batches of rice leaves treated with the freshly purified Xoo cellulase and the buffer

纤维素酶（Cellulase）是水稻白叶枯病菌（Xanthomonas oryzae pv. oryzae，简称Xoo，水稻白叶枯病的致病菌）的II型分泌系统（Type II secretion system）分泌的蛋白，可作为强效诱导剂激活水稻防御反应，包括类过敏反应（hypersensitive response，简称HR）、胼胝质沉积、伴随核碎裂的细胞死亡，并可赋予水稻抵御后续Xoo侵染的功能性抗性。为解析纤维素酶诱导水稻HR的分子事件，本研究采用Affymetrix水稻基因芯片（Affymetrix Rice GeneChips）开展全基因组转录谱分析。
关键词：类过敏反应的表达谱分析
实验设计：将温室培养20天的感病水稻品种台农1号（Taichung Native-1，简称TN-1）的叶片，分别浸润30~40 μl纯化的Xoo纤维素酶（浓度为5 μg/ml），或仅浸润10 mM磷酸钾缓冲液（pH 6.0，实验方法参照Jha等2007年发表于《分子植物-微生物互作》（MPMI）第20卷第31-40页的研究）。处理前48小时将植株转移至生长室，培养条件为28℃、相对湿度80%、12/12小时光暗周期。浸润处理12小时后，采集各处理组中覆盖浸润区域的20~30片叶组织。从混合样本中提取总RNA，采用Affymetrix基因芯片系统开展表达分析。本实验设置3次独立生物学重复，分别使用3批经新鲜纯化的Xoo纤维素酶和缓冲液处理的水稻叶片提取的RNA完成实验。