Sagro-Bacterium-Mediated Transformation Of Potato With Epsps Gene

In this study, EPSPS gene was transferred into potato variety "Favorite"by agro-bacterium-mediated transformation, Microtuber discs were used as explants. The transgenic plants were tested by PCR, PCR-Southern blotting and Southern blotting, and EPSPS gene was confirmed to be integrated in the genome of potato. EPSPS gene was also confirmed to be transcripted by RT-PCR detec tion. An efficient potatoes regeneration and genetic transformation systemsystem were established, at the same time, the influence factors of genetic transformation were also researched such as the kind and concentration. of different phytohormones and bactericides,the time of co-culture, screening methods.The transgenic potato plants were primarily confirmed to be resisted to glyphosate Through identification the resistance to glyphosate. The main results were showed as following:1.The oplimal regeneration medium for tuber disc was MS+1mg/L IAA+4mg/L ZT.2. For EPSPS gene, the optimal concentration of Agrobacterium was OD600=0.5, The optimal time of infection was6minutes. In this condition,the rate of tuber disc with Favorite was72.43%, contamination rate was8.7%.3. Co-culture time had great influence on callus induction rate, The optimal co-culture time of tuber was3days.4. In the comparative experiment of different bactericides, the effective inhibitory concentration of Carbenicillin to agrobacterium was450mg/L, the effective inhibitory concentration of Cefotaxime produced by Haerbin Pharm. Group was450mg/L too. the effective inhibitory concentration of Cefotaxime produced by Invirogen company was150mg/L, and it had no bad effect on young explants under this concentration5. Phosphinothricin was used as the selective agent, The manner of selection was conducted5days later.The concentration was6mg/L at the stage of tuber germination inducing,and4mg/L for the root formation.6.38resistant plants of’Favorite’were obtained andl7plants of them were confirmed to positive by PCR detection.10plants had hybridization signals by further test of PCR-Southern.After that,6plants were confirmed to positive by RT-PCR test, It demonstrated that epsps gene can be transcribed nomally.And these resistand plants were analyzed by Southern-blot,5plants were confirmed to positive. Southern blot showed that single copy of the epsps gene were inserted into the genome of the transgenic plants.7.5transgenic plants of T1generation were tested by PCR,4plants were confirmed to be positive, The detection result of glyphosate resistance showed that the endurance concentration was4.0g/L, It was5-6times to the concentration that used in the field.It is showed that EPSPS gene could be expressed in the progenies.