| Common bean is a kind of important vegetable. It is full of nutriments and tastes delicious. So it has been cultivated all over the world. However, during growth and development,it will be subjected to adverse environmental conditions, such as drought, low temperature and soil salinization and other diseases, which could cause a huge loss to the production of common bean. While in the practical production of common bean, drought is one of the major limiting factors. It is of great significance to identify key regulators in drought response to improve the drought tolerance of common bean. 14-3-3 protein is an important regulation factor for plant response to drought stress. There is no report about response to drought stress in common bean. This research was based on the previous research and preliminary cloning work in our laboratory. In this study, nine 14-3-3 protein genes in common bean have been identified by the methods of bioinformatics. The gene expressions in different tissues and expression patterns under drought stress were analyzed. Then we cloned the promoter region of PvGF14 a gene because its expression level has great change under drought stress. Finally we made a preliminary analysis of drought resistance on transgenic Arabidopsis and tomato, respectively. The main results are as follows:1. In our previous study, according to the genome database from website(http://phytozome.jpi.doe.gov/pz/portal.html), we found nine 14-3-3 protein genes of common bean, and eight of them were cloned. In the present study, we analyzed their protein properties, subcellular localizations, gene structures, homologies and evolutionary relationships with other plant 14-3-3 proteins using bioinformatic method. We found that they had high homology with soybean 14-3-3 gene. Therefore, according to the naming methods in soybean of 14-3-3 proteins, we named these nine genes as PvGF14 a, PvGF14 b, PvGF14 c, PvGF14 d, PvGF14 e, PvGF14 g, PvGF14 h, PvGF14 n, PvGF14 q, which were located in the cytoplasm, the mitochondria and the chloroplast. By analysing the exon and intron of 14-3-3 gene structure in common bean, we found that the exons and introns of 14-3-3 gene which are classified in the same group have the similar size.2. We downloaded the transcription data of 14-3-3 protein gene in different tissues in common bean from phytozome website, and then by the heat map analysis, we found that their expression patterns in common bean were different and isoform specific. And PvGF14 a had the highest expression of transcription in common bean in all kinds of tissues. qRT-PCR results indicated that the transcription expression of 14-3-3 genes in common bean were different when it was subject to drought stress: transcript expression levels of PvGF14 n, PvGF14 c and PvGF14 a were increased; while the PvGF14 e, PvGF14 g and PvGF14 h expression level presented downtrend; and the others PvGF14 d, PvGF14 b transcription was not obvious. While under drought stress the change of PvGF14 a of transcript expression was the largest one.3. We cloned the promoter region of PvGF14 a gene about 1394 bp which was fused with GUS reporter gene and expressed in Arabidopsis. The results indicated that GUS was expressed in seedling, stem, flowers, and silique.4. The result of transgenic Arabidopsis and tomato plants with PvGF14 a gene were obtained. Mannitol simulated drought stress and soil drought stress study indicated that PvGF14 a was involved in drought stress signaling, moreover it could have a negative regulating function.These research results laid a foundation for improved cultivars by further utilizing molecular genetics. |
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