Cloning And Functional Analysis Of Tissue And Organ Specific Promoters From Arabidopsis And Rapeseed

In this research, we obtain five specific promoters information on Arabidopsis thaliana by bioinformatics tool, and find two integrated promoters and one partial promoter in Brassica napus genome by homologous analysis. Based on the analysis, we cloned the promoters and constructed expression vectors including reporter gene for promoter test through transformation. The main results are as follows.1. Cloning and functional analysis of tissue and organ specific promoters from Arabidopsis thalianaAccording to the Arabidopsis thaliana genomic sequence, we designed primers and acquired five specific promoters, they were constructed by vector pBI121, through the Agrobacterium-mediated transformation system, transgenic plants were gained, and the T₀ generation were confirmed by PCR of selective marker gene NPT II and candidate DNA fragment. GUS staining was carried through on T₁ generation to analyze promoters function. The reporter gene was identified only expressed specifically in root, stem, leaf, silique and seed, respectively.2. Effect of CycD3 driven by tissue and organ specific promoters from Arabidopsis thaliana on plant developmentTo understand the effects of Cycd3 under the control of specific promoters on plant development, we prepared three expression vectors. In transgenic plants transformed with the constructs, two of them were observed to have obvious changes in plant development, mainly behaving a delayed growth. Furthermore, expression of Cycd3 under the control of root specific promoter resulted in root bulk diminished and growth lagged. 3. Cloning and functional test of leaf specific promoter from Brassica napusWe identified two integrated promoters in Brassica napus genome by blast tool and transgenic plants were produced by Agrobacterium-mediated transformation. Subsequent functional tests showed that they expressed in the leaf, similar to leaf specific promoter from Arabidopsis thaliana.4. Cloning of flanking sequence of Brassica napus homologous to stem specific promoter from Arabidopsis thalianaA fragment of 1073bp homologous to Arabidopsis stem specific promoter was acquired in Brassica napus genome. And furthermore flanking sequences of 3.4Kb were isolated and compared with stem specific promoter from Arabidopsis thaliana, and two highly homologous regions were identified located in region A, which contains 735bp and the homologous proportion to 74%, and region B, which contains 133bp and to 68%, respectively.