New Strategy to Reconstruct Agrobacterium-mediated Plant Transgenic Expression Vectors

According to the characteristics and limitations of plant transgene vectors,the and transgenic plant expression vectors were reconstructed by cloning,digestion and recombination in the base of the existing commercial vectors.First,design primers and amplify the gene fragments which were composed of CaM35S promoter,gusA gene and NOS terminator three parts from pCAMBIA 1301 plasmid.Recombinate the gusA gene fragments vector into pGM-T to get pGM-35S-GUS-NOS recombinant plasmid by TA cloning.Second,double digeste on cold resistance gene AnGPAT of and the pGM-35S-GUS-NOS plasmid,then connected to get intermediate vector pGM-35S-GPAT-NOS.Third,the intermediate vector and the dual eukaryotic expression vector pCAMBIA 2300 were double digested and connected to get the plasmid p2300-35S-GPAT-NOS.At last,transformed the plasmid p2300-35S-GPAT-NOS into E.coli and Agrobacterium.After PCR and restriction analysis,the transformation plant expression vector p2300-35S-GPAT-NOS was obtained.Experiments show that it is a simple,efficient,universal and economic reconstruction strategy for plant transgene vectors.