Chimeric gene for the transformation of plants

Abstract

Chimeric gene for conferring to plants an increased tolerance to a herbicide having as its target EPSPS comprises, in the direction of transcription, a promoter region, a transit peptide region, a coding sequence for glyphosate tolerance and a polyandenylation signal region, wherein the transit peptide region comprises, in the direction of translation, at least one transit peptide of a plant gene encoding a plastid-localized enzyme and then a second transit peptide of a plant gene encoding, a plastid-localized enzyme. Production of glyphosate-tolerant plants is disclosed.

We claim:  
     
       1. A chimeric gene for conferring to plants an increased tolerance to glyphosate comprising, in the direction of transcription, a promoter region, a DNA sequence encoding a first transit peptide from a ribulose-1,5,-bisphosphate carboxylase small subunit, a DNA sequence encoding an N-terminal domain of a mature ribulose-1,5-bisphosphate carboxylase small subunit, a DNA sequence encoding a second transit peptide from a ribulose-1,5,-bisphosphate carboxylase small subunit, coding sequence for 5-(enolpyruvyl)shikimate-3-phosphate synthase and an untranslated polyadenylation signal. 
     
     
       2. The chimeric gene according to claim  1  wherein the coding sequence for 5-(enolpyruvyl)shikimate-3-phosphate synthase is of bacterial origin. 
     
     
       3. The chimeric gene according to claim  1  wherein the coding sequence for 5-(enolpyruvyl)shikimate-3-phosphate synthase is of plant origin. 
     
     
       4. A vector for transforming plants, which comprises a chimeric gene according to claim  1 . 
     
     
       5. A vector for transforming plants, which comprises a chimeric gene according to claim  2 . 
     
     
       6. A vector for transforming plants, which comprises a chimeric gene according to claim  3 . 
     
     
       7. An Agrobacterium, which contains a vector according to claim  4 . 
     
     
       8. An Agrobacterium, which contains a vector according to claim  5 . 
     
     
       9. An agrobacterium, which contains a vector according to claim  6 . 
     
     
       10. A transformed plant cell, which contains a chimeric gene according to claim  1 . 
     
     
       11. A transformed plant cell, which contains a chimeric gene according to claim  2 . 
     
     
       12. A transformed plant cell, which contains a chimeric gene according to claim  3 . 
     
     
       13. A transformed plant with improved glyphosate tolerance, which was obtained by regeneration of the cell according to claim  10 . 
     
     
       14. A transformed plant with improved glyphosate tolerance, which was obtained by regeneration of the cell according to claim  11 . 
     
     
       15. A transformed plant with improved glyphosate tolerance, which was obtained by regeneration of the cell according to claim  12 . 
     
     
       16. A plant according to claim  13 , which is a dicotyledon. 
     
     
       17. A plant according to claim  13 , which is a monocotyledon. 
     
     
       18. A process for constructing a chimeric gene according to claim  1 , wherein sequences for at least two transit peptide regions, at least one sequence of the N-terminal domain of a mature ribulose-1,5-bisphosphate carboxylase small subunit, at least one sequence encoding 5-(enolpyruvyl) shikimate-3phosphate synthase and a polyadenylation signal region are each isolated, and wherein said sequences are then assembled in the direction of transcription of the 5-(enolpyruvyl) shikimate-3phosphate synthase gene. 
     
     
       19. A process for constructing a chimeric gene according to claim  2 , wherein sequences for at least two transit peptide regions, at least one sequence of the N-terminal domain of a mature ribulose-1,5-bisphosphate carboxylase small subunit, at least one sequence encoding 5-(enolpyruvyl) shikimate-3-phosphate synthase and a polyadenylation signal region are each isolated, and wherein said sequences are then assembled in the direction of transcription of the 5-(enolpyruvyl) shikimate-3-phosphate synthase gene. 
     
     
       20. A process for constructing a chimeric gene according to claim  3 , wherein sequences for at least two transit peptide regions, at least one sequence of the N-terminal domain of a mature ribulose-1,5-bisphosphate carboxylase small subunit, at least one sequence encoding 5-(enolpyruvyl) shikimate-3-phosphate synthase and a polyadenylation signal region are each isolated, and wherein said sequences are then assembled in the direction of transcription of the 5-(enolpyruvyl) shikimate-3-phosphate synthase gene. 
     
     
       21. An agronomic method comprising: 
         A )  growing a plant which contains in its genome a nucleic acid sequence encoding a polypeptide sufficient for localization of a gene product in a chloroplast of a plant cell, which polypeptide comprises in the direction of translation:    
       ( i )  a first chloroplast transit peptide of a ribulose -   1 , 5   - bisphosphate carboxylase oxygenase small subunit;    
       ( ii )  an N - terminal domain from a mature ribulose -   1 , 5   - bisphosphate carboxylase oxygenase small subunit; and    
       ( iii )  a second chloroplast transit peptide of a ribulose -   1 , 5   - bisphosphate carboxylase oxygenase small subunit;    
         and further encoding a  5   -( enolpyruvyl ) shikimate -   3   - phosphate synthase which renders said plant tolerant to a phosphonomethyl glycine herbicide; and    
         B )  applying said herbicide to the plant.   
     
     
       22. The method of claim  21 , wherein the plant is maize. 
     
     
       23. The method of claim  21 , wherein the  5 -( enolpyruvyl ) shikimate -   3   - phosphate synthase is mutated, the mutation being selected from the group consisting of Pro  101  to Ser and Gly  96  to Ala.   
     
     
       24. The method of claim  23 , wherein the plant is maize.