تاثير عناصر ريزمغذي هاي آهن و روي بر روي عملكرد و اجزا عملكرد كشت دوم دو رقم آفتابگردان در منطقه ارسنجان

Isolation,molecular analysis and site directed mutagenesis in E. coli EPSPS gene in order to make glyphosate tolerant rapeseed (Brassica napus L.)

The manipulation of bacterial EPSPS gene in order to reduce its affinity to glyphosate, is one of the most effective methods for production of glyphosate tolerant plants. In this research, we study on glycine 96 of E. coli EPSPS enzyme. This amino acid is an important residue for EPSPS-Glyphosate binding.We used site directed mutagenesis method to inducing a point mutation in E. coli EPSPS gene to convert glycine 96 to alanine. The manipulated EPSPS gene was cloned in pUC18 as an universal cloning and pBI121 as a plant expression vectors. The results of molecular analysis and sequencing showed that the manipulated genes has been changed and cloned in correct orientation in both plasmids. Recombinant pBI121 containing altered EPSPS gene were transferred to rapeseed (B. napus), PF-7045-91 cv, via Agrobacterium tumefaciens mediated transformation method. The target tissue for transformation was the cut end of cotyledonary petiols. At the first step the transformed explants were screened in kanamycin containing media. Result showed that plant regeneration and transformation frequency was 73 and 30% respectively.Glyphosate tolerance will be assayed in next generation.