Cloning And Functional Analysis On Proline Transporter Genes Gmprot1and Gmprot2from Soybean

Enviromental stresses, such as drought and high salinity have adverse effects on plant growth, development of agricultural and quality of production. To adapt such detrimental conditions,plants have developed a variety of defense strategies,requiring many genes regulated by abiotic stress.Although a large number of genes regulated by salt and drought stress have been identified and verified their function in abiotic stress,the biological functions of many genes related to abiotic stress are still largely unknown in higher plants. In this study, we cloned GmProTl and GmPorT2genes were isolated from soybean;studyed on sequence and protein structures of GmProTl and GmProT2with bioinformatics; analysed expression pattern of GmProTl and GmProT2with Real-time RT-PCR;.studied on functional analysis of transgenic GmProT1and GmProT2gene Arabidopsis which were transformed via Agrobacterium mediation after treated with various concentrations of exogenous Pro,drought and high salinity, in order to study the contribution of the genes in plant to stress tolerance system,and provide a theoretical basis for studying on resistance soybean gene of Stress response in the reaction mechanism. The results are as follows:1. Nucleotide sequence of AtProT1was used to search soybean gene bank for high homology soybean gene sequence. GmProT1and GmPorT2genes were isolated from soybean. The phylogenetic relationship appeared that GmProTl and GmPorT2genes were more similar to Populus trichocarpa ProT and Arabidopsis ProT gene than others. GmProTl and GmProT2genes consist of11membrane-spanning regions.2. Investigate on the genes expression patterns of multiple tissues in soybean found that GmProTl gene expression of root was highest at seedling stage; that of stem and leaf were highest at bearing pod stage; that of flower was highest at flowering stage. GmProT2gene expression of root, stem, leaf and flower were highest at bearing pod stage.GmProTl and GmProT2genes expression of seed were increased at seed development stage from20DAF (seed of20days after flowering), until reached at the highest level on35DAF.3. The specific expression pattern of soybean GmProT1and GmProT2induced by stresses indicated that the GmProT1and GmProT2genes expression of leaf were the highest; root of GmProT1and GmProT2genes were most sensitive and reached peak value earliest.4. After transformed GmProT1and GmProT2genes into Arabidopsis via Agrobacterium mediation, transgenic Arabidopsis.After Arabidopsis were treated with various concentrations of exogenous Pro,we found that germination rate of seeds and root elongation increased at5mM exogenous proline,transgenic Arabidopsis were significantly higher than wild Arabidopsis.While at50mM or100mM exogenous proline, transgenic Arabidopsis were obviously lower than wild Arabidopsis. Moreover, proline content in transgenic Arabidopsis was significantly higher than wild Arabidopsis. After they were treated by high salt and drought stresses, germination rate of seeds and root elongation at seedling stage or at cotyledon stage of transgenic Arabidopsis were significantly higher than those of wild Arabidopsis. Applied exogenous5mM proline when Arabidopsis were treated by high salt and drought stresses, it increased seed germination rate and root elongation of Arabidopsis. It relieved stresses hurt to plants.Transgenic Arabidopsis were significantly better than wild Arabidopsis. Proline content in transgenic Arabidopsis was significantly higher than wild Arabidopsis at adult stage after treated with hight salt and drought stresses. Above all, under the same condition, transgenic Arabidopsis transported more proline than wild Arabidopsis, and they increased observably salt and drought tolerance of Arabidopsis.