One of the important factors leading to osmotolerance (drought and salinity) is accumulation of osmoprotectors such as amino acids: Proline, glyicine and glycine betaine; compounds like manitol and pinitol and also HSP proteins by cells of living organisms. The biosynthetic pathways of these products are similar in bacteria and higher plants, therefor we can transfer the genes responsible in production of these compounds to plants in order to obtain transgenic lines tolerant to osmoticstress. In this project we used of gamma radiation which is monochonic and has selective penetrancy to produce mutant strains of Escherichia coli strain MK 148. Salt tolerant colonies were obtained and isolated. The strains were able to tolerate 600 millimolar salt in the media and have normal growth and reproduction; It must have been mutations in gene coding glutamyl kinase (proB) which is the first enzyme in the proline pathway. After this mution, the sensitivity of glutamyt kinase to End-product feedback inhibition has been reduced. The genes of glutamyl Kinase (proB) and glutamic semi- aldehyde dehydrogenase (proA) were isolated by polymerase chain reaction (PCR) from salt- tolerant colonies and cloned in PUC 19. This plasmid was transformed to bacteria, which were unable to biosynthese proline to make them to produce proline as well as to growth in salty media.