Identification For Cis-acting Elements Of Promoter Of The Gene Sm-Nvas Related To Vascular Bundle Development In Nicotiana Tobacum

The promoter plays a key role in genetic engineering,which determines the transcription pattern of exogenous genes.It is important for biological research on an appropriate promoter to regulate the expression of different exogenous genes.Tissue-specific promoter can drive exogenous gene to express efficiently in the specific parts of the plant,which can avoid wasting the nutriment as the constitutive promoter.The focus of the promoter study is to explore the appropriate and efficient tissue-specific promoters and to analyze their cis acting elements.The sequence of sm-Nvas promoter was cloned from W38,wild type in Nicotiana tobacum,which was verified involved in the development of vascular bundle by bioinformatics analysis and transgenic experiment in the previous study.This study aimed to analyze the drive activity of the promoter and to identify the cis-acting elements of the promoter.The main results of this study are as follows:(1)The method of 5' end deletion analysis was used in this study.A of initiation codon ATG as the starting site +1,the sm-Nvas promoter region(-799 ? 0)was divided into 20 fragments of different lengths at 5' end region.20 fragment-GUS recombinant vectors and 20 fragment-GFP recombinant vectors were constructed.35 S promoter on p CAMBIA1301 and p CAMBIA1302 were replaced by the above 5'end deletion fragments,respectively.Transgenic plants were obtained by Agrobacterium tumefaciens mediated leaf disc transformation method.(2)Based on GUS staining and GFP observation,it showed that sm-Nvas promoter belongs to a tissue-specific promoter.The expression of the reporter genes were detected in all of the transgenic plants.The-216 region of 5' UTR region was sufficient to drive the exogenous gene to express in vascular bundle.It suggested that the-216 region contains the core element of the promoter and the element of vascular bundle specific expression.(3)The drive activity of sm-Nvas promoter was much higher than that of the 35 S promoter.The sm-Nvas promoter was a vascular bundle specific promoter with strongactivity.The expression of reporter gene of transgenic plants containing-402 ?-640 region was significantly increased,and-640 region had the highest activity.The results suggested that there was an element enhancing promoter activity in-402 ?-337 region.And an element existed in-680 ?-640 region,which could inhibit the activity of the promoter.