ABSTRACT: To comprehend the gene expression profile in rice flag leaf under high temperature, Agilent 4×44k rice oligo microarray experiments were carried out using rice flag leaf of reproductive stage at 0 min, 20 min, 60 min, 2 hr, 4 hr, and 8 hr after the treatment of 40 degree centigrade, and the significantly expressed genes mainly involved in transcriptional regulation, transport, protein binding, antioxidant, and stress response were identified. Among them, the predominant transcription factor gene families were Hsf, NAC, AP2/ERF, WRKY, MYB, and C2H2. KMC analysis discovered the time-dependent gene expression pattern under heat. The metabolism pathway analysis demonstrated that, under heat treatment, glycolysis and ubiquitin-proteasome was enhanced, and TCA, gluconeogenesis, the secondary metabolism and light-reaction in the photosystem was dramatically repressed, which revealed the great importance of maintaining primary metabolism and protein homeostasis in response to heat in rice flag leaf.