Study On The Promoter Of RSSG8 Gene In Rice(Oryza Sative L.)

The promoter of RSSG8 gene in rice was cloned through twice genome walking and it was fully sequenced .This sequence was named as R8PN.Comparing R8PN sequence and RSSG8 gene with nr database of GenBank in NCBI, the Blastn results indicated that R8PN and RSSG8 are located in the NO. 12 chromosome of Oryza sativa genome (Japonica .culticar-group, accession number: AL772420.6 and AL772425.7), and they are close in sequence. Moreover, R8PN and RSSG8 were searched in the contig Scaffold2153 by comparing with Oryza sativa genome (indica culticar-group) from SCBI (Sichuan University Bioinformatics Centre, http://scbi.scu.edu.cn/), and they are close in sequence. The Net servers PLACE (http://www.dna.affrc.go.jp/htdocs/PLACE/) and PLACE CARE (http://sphinx.rug.ac.be:8080/PlantCARE/) were used to analyze the motif of promoter. The final results indicated that the traditional TATA-box was not found in the R8PN sequence, suggesting that the region (-33-46) enriching A and T could be an especial RNA polymerase binding site. Several CAAT-box locate in R8PN, suggesting that its play an important role in the expression of RSSG8 gene. Moreover, an abundant class of light responsive element such as G-box, GT1-motif,GATA-box, CG-motif, GAG-motif and LAMP-element exit in R8PN sequence. There are six G-boxes, two GATA-boxes and GAG-motifs, which suggest that the expression of RSSG8 is regulated by light.In order to identify the key functional regions of the promoter, the authors insert R8PN's two deletion sequences into another vector pBI121 to take place of CaMV 35S promoter to get two expression vectors pRGUS1 and pRGUS2. Re-combinant plasmids were transformed into Agrobacterium tumefaciens EH 105,and the later were used to infect leaves and mature pollen of tobacco.GUS transient expression demonstrated that both deletion fragments could work as promoter,but their efficiency were different from each other in mature pollen of tobacco. The efficiency of the 260bp deletion fragment is obviously better than the 471bp deletion fragment, which may suggest that there are negutive regulative-factors in the -261-471 region of R8PN sequence.