Transformation Of Phytase Gene Into Soybean

Soybean is an important economic crop,. Soybean is playing an important role in the production and national economy. At present, the application of fertilizer in a large number of phosphorus and inositol synthesis of soybean roots in the soil is difficult to absorb and utilize of phytate on soybean growth and development, cultivation and production have had a great influence. However, this method not only improves the cost of planting, resulting in extreme waste of phosphorus pollution of the environment by increasing P fertilizer can play the utilization of soybean to phosphorus. Increase of P fertilizer is not an effective way. However, with the tremendous development of transgenic technology matures and a wide range of applications, we can by transgenic technology, the soybean root-specific promoter promoter gene connected with exogenous phytase gene expression vector, imported soybean to plant the acid gene expression in soybean roots, the roots of the expression product of phytase can be broken down into the soil of phytate phosphorus conducive to the absorption of soybean, soybean on soil phosphorus utilization to improve soybean yield reduce planting costs, to provide an effective way to provide a theoretical basis for improving the quality of the soybean, but also reduce the pollution caused by the environment.In this study, we use the soybean rootspecific promoter gene (RSP) to connect to the pCAMBIA3300-PhyA-bar expression vector which had constructed, then transformation. The recombinant expression vector transformed by Agrobacterium-mediated and pollen tube pathwayexogenous gene into the soybean. In order to get phytase gene only in the root-specific expression of genetically modified soybeans. The results are as follows:(1)Extract the roots of specific promoter (RSP) from AHLG plasmid, according to AHLG designed two pairs of specific primers and PCR amplification. In order to get rootspecific promoter (RSP) gene fragment. Then we use DNAman software analysis of the pCAMBIA3300-PhyA-bar and root-specific promoter (RSP) restriction enzyme sites, analyze the results pCAMBIA3300-PhyA-bar and root-specific promoter (RSP) with restriction enzymes PstI and Xbal enzyme cut, the fragment and the plasmid without undermining the purpose. Connection the PCAMBIA3300-PhyA-bar roots-specific promoter (RSP), and get the recombinant expression vector pCAMBIA3300-RSP-of PhyA-bar.(2) Select four genotypes of soybeans was transferred pCAMBIA3300-RSP-PhyA-bar recombinant vector by embryonic tips and cotyledonary node. four genotypes of soybean embryonic tips for basic research into the different genotypes in6-BA and IBA, the sensitivity of the optimization of culture conditions Determine the concentration of four genotypes optimal6-BA and IBA. Transformed plant regeneration PCR testing, we get a positive result of plant. Initial proof of the expression vector has been transformed into successed.(3) We transferred recombinant expression vector pCAMBIA3300-RSP-PhyA-bar into soybean by pollen tube path way, harvest seeds and let it grow,then extract genomic detection with PCR, access to a test positive for the plant pollen tube pathway. Initial proof of the expression vector has been transformed into success.(4)We use PCR RT-PCR, Southern-Blot analysis T1. In order to identification recombinant expression vector could expression and stably heredity to offspring. Taking test positive for the T1for enzymatic activity.determination the condition of PhyA.