Gene Expression Profile Under Salt Stress And Function Analysis Of Salt Respond Tanscription Factor Genes Bzip110, Wrky49and Wrky111in Soybean

There are about30million hectares of salinization land in China. Therefore soil salinization becomes one of the main limiting factors in agricultural production. Considerably, researches on the mechanism of salt tolerance, selection and development of salt tolerant varieties have profound significances for the improvement ecosystem in salinization area and the sustainable application of land resources.In this regard, soybean is an important industrial crop in the world. To research the mechanism of salt tolerance in soybean can be helpful for the genetic improvement for salt tolerance in soybean. In this study, we analyzed differentially expressed genes under salt stress in coastal wild soybean (Glycine soja) and cultivated soybean (Glycine max) by using DGEP (digital gene expression profiling) technology. Three differential expressed genes, bZIP110, WRKY49and WRKY111were selected as the candidates of salt response genes for further study on their physiological function and molecular mechanism.In DGEP data, there were1327genes which responded salt stress in Glycine soja line "Yedade2",826up-regulated,501down-regulated. Our results demonstrated there were3627genes responded salt stress in Glycine max cultivar "Dongnong690",1709up-regulated,1918down-regulated. We got the same result with DGEP data about some respond genes’expression tendency using qRT-PCR. In addition, we chose significant salt respond genes as candidates including bZIP110, WRKY49and WRKY111, to research their function on salt stress using composite plants and transgenic plants. In the result we found that all the candidate genes could improve salt tolerance of composite and transgenic plants.Transcription factor gene bZIP110is1542bp in soybean genome, with no intron, coding168amino acids. Its subcellular localization is in the nucleus and belongs to S group. bZIP110was induced by200mM NaCl stress, responded salt stress distinctly in Glycine soja and Glycine max under200mM NaCl for12h. In the analysis of expression in different tissues, we found it had a high expression level in the root, stem and leaf in Glycine max, while had a low expression level in leaf only in R4(full pod stage). Moreover our results indicated that, bZIP110enhanced salt tolerance in transgenic Arabidopsis plants, the content of Na+was less than in transgenic plants’leaves, and the expression level of stress respond genes MYB2, PAD3, UGT71B, LCL1, DREB2, NHX1, SOS1and a POD (peroxidase) gene RCI3was higher than wild type. POD can remove H2O2(hydrogen peroxide) and the toxic of phonel, keep oxidative balance in plant under abiotic stress, and this may related to the major reason of more salt tolerance of transgenic plant than wild tpye. The expression levels of genes CCA1、LTP3and P5CS were no difference in transgenic plant compared to wild type, but the expression level of LHY was down regulated. Considerably we realized that, bZIP110had an activation function and interactive role with bZIP105belonging to C group in Y2H (yeast two hybrid), and could bind ACGT motif in Y1H (yeast one hybrid). But we must mention that, the capacity of binding ACGT motif was recorded weaker than bZIP105. We also infer that bZIP105binds promoter of a gene and bZIPl10activates it under salt stress in plant.Transcription factor gene WRKY49is3519bp in soybean genome, with4introns, coding575amino acids. Its subcellular localization is in the nucleus. It has two WRKY domains and one zinc-finger motif, belongs to group Ⅰ. WRKY49was induced by200mM NaCl stress, it responded salt stress more swift and violent in Glycine max under200mM NaCl, but the respond was more stable in Glycine soja, this maybe has relations with the reason of Glycine soja more salt tolerant. WRKY49expressed in all tissues,it had a high expression level in root and leaf, low expression level in pod. Over-expression WRKY49could enhance the salt tolerance in transgenic tobacco and Arabidopsis plants. Germination rate of seeds was lower and length of root was shorter in Arabidopsis mutants than wild type under salt stress. The POD gene RCI3was significant up regulated in transgenic Arabidopsis, stress respond genes LHY, UGT71B6, DREB2, PAD3, LTP3, HNX1and SOS1were up regulated too, but the content of Na+in transgenic leaves was no distinct difference compared to wild type. So we conclude that WRKY49have the ability to enhance the compartmentation of Na+in transgenic plant.Transcription factor gene WRKY111is3618bp in soybean genome, with3introns, coding398amino acids. It has one WRKY domains and one zinc-finger motif, belongs to group Ⅱc. WRKY111was induced by200mM NaCl stress, it responded salt stress more swift and violent in Glycine max under200mM NaCl, but the respond was more stable in Glycine soja, this maybe has relations with the reason of Glycine soja more salt tolerant. WRKYlll mainly expressed in root, low expression level in leaf and flower, no expression in pod. Over-expression WRKY111could enhance the salt tolerance in transgenic tobacco and Arabidopsis plants. The expression level of POD gene RCI3was100folds higher in transgenic Arabidopsis than wild type, and also up-regulated other six stress-respond genes DREB2, MYB2, PADS, LTP3, NHX1and SOS1. Four genes CCA1, LCL1, UGT71B6and P5CS expression levels in transgenic plants were no differences with wild type, but the expression level of LHY was down regulated. The content of Na+was less in transgenic plants’leaves than in wild type. Moreover, we confer that the major reason of WRKYlll enhancing salt tolerance in transgenic plants is up-regulating RCI3and limiting the accumulation of Na+in leaves.