Promoter Analysis And The Effect On Stress Response Of Gene BRNO3 In Arabidopsis Thaliana

BRN03 is an unknown gene in Arabidopsis. It is located on chromosome IV of Arabidopsis thaliana. In order to study the biological function of BRNO3, we study the promoter sequence and CDS regions of BRNO3 in this pape.We make the bioinformatics analysis of BRNO3. BRNO3 has no trans-membrane domain and signal peptide. We predicted its tissue expression, finding that it expresses in all tissue, but in addition to inflorescences, sepals and folders, its expression level is very high, the expression of BRNO3 in other tissues is very low. In addition to the BRNO3 secondary and tertiary structures were predicted and found that protein structure has PD40 repeats, TolB domain, and DPP-IV domain. PD40 has a similarity with the WD40.RT-PCR results showed that after cold and penetration stress treatment, the expression of BRNO3 in green tissues and roots was enhanced. After heat stress treatment, the expression of BRNO3 was decreased both in the green groups and root. After salt stress treatment, the expression of BRNO3 in green tissues was enhanced, while there was no affect in root.To further study the tissue expression specificity of BRNO3, we cloned the BRNO3 upstream length 1030bp (P1) and 386bp (P2) two sequences as the promoter region. The P1 1030bp fragment contains the BRNO3 promoter sequence and part of its upstream coding sequence, while the P2 386bp fragment contains only the promoter coding sequence. Gene Transfer and GUS expression analysis showed that the expression of P2 promoter sequence fused GUS (P2-GUS) is more stronger than P1 promoter fusion GUS (P1-GUS).However, the expression of P2-GUS is not regulated by cold, heat, salt, osmotic stress and brassinolide; while P1-GUS response to various stresses and BR. These results suggest that BRNO3 adjacent upstream gene 3'fragment play the important role in regulating the expression of BRN03.P1-GUS expression analysis showed that under non-stress the main part of GUS expression is leaf, the growing point of stem and the mature region while there is no expression in the petiole, stem and the tip of root. Stress led to the GUS expression were outside most of the tissue in the Arabidopsis seedling, and the expression increased. These results indicate that BRN03 response to variety of stress. It is a stress-related gene. It may has an important role in stress resistance in Arabidopsis. Therefore, we also built the BRNO3 YFP fusion over expression vector for further study of the gene function has laid a solid foundation.