Identification And Functional Characterization Of Pathogen-inducible Regulatory Elements Of Promoter Pgrmzm2G084947for R.Solani Caused Maize Sheath Blight Disease On Maize

Maize sheath blight were becoming the serious and major diseases in mian production region of world. However, there was little research in molecular mechanism and disease resistance path was not clear. It was an important limitation for breeding resistance variety and disease control without major resistance gene identified in germplasm resources. Therefore, it is essential for exploring actively the maize sheath blight resistance path for disease control. Previous studies have shown that cloning of the maize disease resistance gene and the promoter can have a thorough understanding of the host and pathogen interaction, which can lay the foundation for genetic engineering improvement. Through the digital gene expression profiling (RNA-seq) between maize and different virulent strains YWK62and YWK196about300differential expression genes was identified. In this study, one up-regulation gene, GRMZM2G084947was selected to validate the function of promoter, and to identify the czs-elements which response to Maize sheath blight pathogen.About2kb DNA fragment upstream the translation initiation site was cloned into the promoter expression vector by PCR technology which contains the reporter gene GUS. Then1391::pGRMZM2G084947transgenic individuals was generated with Agrobacterium-medlated transformation in rice variety Zhonghua11. It found that this promoter can generate inducing response to maize sheath blight fungus YWK196quickly and strongly, and has very high expression in younger leaves and plant stem. According to promoter prediction software PLACE and tobacco transient expression system, we detected report gene expression and analysed the truncated promoter fragment.Firstly, the key regions of promoter induced by the pathogenic fungus was identified:-1144bp to-1084bp and-1084bp to-973bp. Through PLACE analysis found that one region (-1084bp to-973bp) contains the GT-1element,which is a czs-element associated with pathogen and salt response.Another regional144bp to-1084bp) found there was no known promoter element. According to truncating analysis for this unknown region, we determined the promoter core region in-1122bp to-1112bp region finally. Through bioinformatics analysis, there is a GGGCA sequence fragments in the10bp interval. This five bases fragments and10bp fragments (-1122bp to-1112bp) which trigger the report gene GFP expression level are about same. However, the full length of the promoter pGRMZM2G084947which trigger the report gene GFP expression levels is about two times of the element GGGCA’s. So the new element GGGCA plays a key role in promoter responding to pathogen.In conclusion, this study identified a new pathogen-inducible element GGGCA in the promoter pGRMZM2G084947which responses to the pathogen of maize sheath blight. The pathogen-inducible promoter and its key cis-element can be applied in plant transgenic breeding to resist to maize sheath blight.