Cloning And Analysis Of Rice Gene Promoter Ospz4

Promoter plays important roles in gene function and regulation, The study of which is helpful to understand the complexity of gene regulation. Rice in the world is one of the most important food crops and the model plants. To isolate and explore the endogenous promoters of rice, the Affymetrix Gene Chip Rice was used to analyze genome expression profiles of different tissues and organs of rice Peiai 64S (Oryza saliva L.) at different developmental stages under drough, cold and heat stress conditions. We has screened a high expression levels Gene OsSG4 (GenBank accession:AK.068991.1) induced under normal and stress conditions.The 1,625 bp upstream promoter region of gene OsSG4 was cloned from the genome of rice Nipponbare. Four subfragments of Ospz4 with different sizes of 1,293 bp,1,006 bp,834 bp and 492 bp were obtained by deletion from its 5'end, each of which was fused with ?-glucuronidas (GUS) reporter gene to form a plant expression vector. The fusion genes were tested for transient GUS expression in tobacco leaves by injection of Agrobacterium cells harboring these gene vectors. The results showed that promoter Ospz4 and its 5' deletion fragments could drive GUS gene expression, The highest GUS activity detected was driven by the 492 bp 5'deletion fragment of Ospz4. For stable transformation, all these vectors were delivered into rice variety Taibei 309 through Agrobacterium-mediated transformation. Histochemical staining and quantitative real-time fluorescent PCR showed that promoter Ospz4 could drive the GUS gene expression in leaves, roots, stems, spikelet, endosperms and coleoptiles of transgenic plants, whose expression activity reached 49% of the double 35S promoter. Similarily, all the 5'deletion fragments could drive the GUS gene to express in the leaves, roots, stems, spikelet and coleoptiles of transgenic plants. The promoter activity of 492 bp fragments in rice leaves was basically the same as the total length of Ospz4, but the activities of the 1,293 bp,1,006 bp and 834 bp fragments were significantly weaker than that of the full-length fragment, reaching 30%,28% and 23% of the double 35S promoter activity, respectively. Under drought, low temperature and high temperature treatment conditions, promoter Ospz4 could maintain a high activity in T1 plant, with the expression levels of 112%,94% and 105% of the normal condition, respectively. In this study, a rice endogenous promoter with constitutive expression was successfully cloned and analyzed. Promoter Ospz4 functions as a constitutive promoter and could drive the GUS gene to express in leaves, roots, stems, spikelet and coleoptiles of transgenic plants. Ospz4 is useful for both scientific research and crop improvement.