Genome-wide Gene Expression Profiling Of Rice Under Heat Stress And Functional Characterization Of Specific Promoters

As one of the most important crops in the world, rice has been encountering varous adverse environmental factors such as heat, drought, disease, which has attracted more and more attentions all around the world. Consequently, it becomes an important task to improve rice resistence to various stresses based on gene engineering via cloning and characterizing stress-related genes and promoters.The expression profile in rice panicle and flag leaf under heat stress was analyzed through rice microarray. Furthermore, using transgenic technology, the expression activity of the three promoters from the highly heat-inducible genes in rice panicle and flag leaf under heat and drought was detected. The major results are as followed:(1) The differentially expressed genes (DEGs) in rice panicle and flag leaf under heat treatment were obtained under the condition of quality control and evaluation using HCL. The analysis on the DEGs showed the time point2h of heat treatment may be an important transition for rice response to heat stress. The results from GO retrieve and gene expression pattern analysis showed the heat-responsive (HR) genes in rice panicle and flag leaf are mainly involved in transporter, protein binding, antioxidation, catalysis, transcription regulation and stress response. The comprehensive statement and primary comparison on the HR transcription factors such as Hsf, AP2/ERF, bHLH, bZIP, Myb, WRKY, NAC and C2H2as well as Hsp family genes from both tissues was carried out. These work provided important clues for the characterization and identification of tissue-specific heat-responsive genes and promoters.(2) The expression profile analysis showed that the complicated changes of ROS-related genes in rice panicle under heat stress helps maintain ROS balance, and that the genes related to auxin, ABA and ethylene metabolism and signal transduction were mainly down-regulated, and the ones involved in light, calcium and G-protein were up-regulated.(3) The analysis on the promoters of co-regulated genes in rice panicle showed that GCC box, CE3, ABRE and HSE are significantly enriched in the early-regulated cluster, and GCC box and CE3are the centers of the two co-occurrence network, with which HSE and ABRE significantly co-occured, respectively.(4) A regulation model central to ROS was established, and the data from the rice panicle microarray and ROS quantification help elucidate the importance of ROS balance in the rice panicle response to heat shock.(5) The pathway analysis on the HR genes in rice flag leaf showed that heat shock significantly promoted glycolysis and UPS, and repressed TCA, secondary metabolism, and photosynthesis via inhibiting light reaction.(6) Based on the screening microarray data, qRT-PCR idenfitication and bioinformatics analysis on promoters, the three candidate promoters for the highly heat-inducible genes were cloned, transformed into rice, and the positive transgenic lines were characterized using PCR. After heat and drought treatment on the mature transgenic rice, GUS enzyme activity and expression level was detected using histochemical staining, fluorescence quantification and qRT-PCR in rice panicle and flag leaf. The results showed that, among the three candidate promoters, Hsfp exhibited highest activity in rice panicle under heat and drought, bu none in flag leaf; Dehp had no activity in both tissues under stress conditions; Putp showed higher activity only in panicle under drought.