Transformation Of Phytoene Synthase Into Inbed Lines Of Zea Mays L. By Agrobacterium

The vitamin A dificiency is listed as one of the world's four major nutrirional dificiency diseases by the World Health Organization.However,as one of the most widely distributed food crops in the world, maize dose not contain vitamin A, which is bound to lead to severe lack of vitamin A in some less developed area where maize is still the stable food. Therefore, the improvement of vitamin A in maize will greatly improve the nutritional status of people in poor areas .Transgenic approach to improve vitamin A content of maize is an economical and practical way.In this thesis,we used Agrobacterium-mediated approach to transform the key enzyme of the carotenoid biosynehesis called PSY(phytoene synthase) into the maize,and we hope to obtain the transgenic maize high of vitamin A. The target gene PSY comes from Eustoma grandiflorum,and was inserted into the plant expression vector pCAMBIA2300 with the 35S promoter and the bar selection marker.The vector was then transformed into Agrobacterium tumefaciens C58.We induced embryonic callus from immature embryos of different inbred lines of maize such as 7922,238,265,p138,etc.,and studied the factors affecting callus induction rates.The results show that when the size of immature embryo is in 1~2mm,and the inoculated immature embryos placed scutellum up,we could obtain a higher callus induction rate.The genotype and medium compositon can influnce callus indution rate significantly.The second type callus of 7922,238 and 265 can be long-term subcultured and regenerate into plants.Medium containing cysteine and silver nitrate can significantly reduced the browning rate. Before the transformation,we studied the resistance of the callus and found that when PPT(L-phosphiothricin) concentration was 5mg/L,80% of the callus died,so the screening concentration is 5mg/L.We also studied the factors affecting Agrobacterium-mediated transformation,and found that when the infection time was 15min,and the co-culture medium was added with glucose and MES,we could get the highest rate of resistant callus.We obtained 17 resistant plants through the selection of PPT,the result of PCR molecular detection indicated that PSY has been inserted into three plants.