Transgenic Plants Resistant To Non-Protein Amino Acids

Abstract

Transgenic plants resistant to bio-herbicides, particularly to phytotoxic non-protein amino acids including the meta-tyrosine (m-tyrosine) amino acid analog and salts thereof, means and methods for producing the transgenic plants.

What is claimed is: 
     
         1 . A transgenic plant comprising at least one cell comprising at least one exogenous polynucleotide encoding an aminoacyl tRNA synthase (aaRS) or a fragment thereof, the aaRS or fragment thereof comprising an editing module capable of hydrolyzing tRNA misacylated with non-protein amino acid analog, wherein the plant is resistant to the non-protein amino acid analog and salts thereof. 
     
     
         2 . The transgenic plant of  claim 1 , wherein said plant grows in a medium containing the non-protein amino acid or salt thereof in a concentration that significantly inhibits the growth of a corresponding non-transgenic plant. 
     
     
         3 . The transgenic plant of  claim 3 , wherein growth inhibition is shown by at least one of reduced root length, reduced root radical, reduced root mass, reduced plant height, aberrant change in a plant tissue morphology or color, reduced plant shoot mass, reduced plant shoot number and any combination thereof. 
     
     
         4 . The transgenic plant of  claim 1 , wherein the non-protein amino acid analog is meta-tyrosine (m-Tyr) compound and the aaRS is phenylalanyl-tRNA synthetase (PheRS). 
     
     
         5 . The transgenic plant of  claim 4 , wherein the m-Tyr compound has a formula of Formula I or a salt thereof, Formula I having the following formula: 
       
         
           
           
               
               
           
         
         wherein: 
         R 1  and R 2  are independently selected from the group consisting of H, sulfonate, sulfonamide, phosphonate, alkyl, alkenyl, alkynyl, alkoxy, alkoxycarbonyl, saccharide, cycloalkyl, heterocycloalkyl, aryl and heteroaryl, wherein each of the phosphonate, alkyl, alkenyl, alkynyl, alkoxy, alkoxycarbonyl, saccharide, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl is either substituted or unsubstituted; 
         R 3  is selected from the group consisting of H, alkyl, alkenyl, alkynyl, alkoxy, alkoxycarbonyl, saccharide, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl, wherein each of the alkyl, alkenyl, alkynyl, alkoxy, alkoxycarbonyl, saccharide, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl is either substituted or unsubstituted; 
         X is selected from the group consisting of O and N—Y, wherein Y is selected from the group consisting of H, alkyl, alkenyl, alkynyl, alkoxy, alkoxycarbonyl, saccharide, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl, wherein each of the alkyl, alkenyl, alkynyl, alkoxy, alkoxycarbonyl, saccharide, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl is either substituted or unsubstituted; 
         R 4 , R 5 , R 6 , and R 7  are independently selected from the group consisting of H, hydroxyl, halogen, amino, and nitro; and 
         R 8  and R 9  are independently selected from the group consisting of H, hydroxyl, halogen, amino, methyl, and halogenated methyl. 
       
     
     
         6 . The transgenic plant of  claim 5 , wherein the m-Tyr compound has a formula of Formula II, Formula II having the following formula: 
       
         
           
           
               
               
           
         
       
     
     
         7 . The transgenic plant of  claim 4 , wherein the PheRS is a heterotetrameric bacterial PheRS composed of two PheRS-α and two PheRS-β strands. 
     
     
         8 . The transgenic plant of  claim 7 , wherein the bacterial PheRS is selected from the group consisting of  Escherichia coli  ( E. coli ) PheRS and  Thermus thermophilus  PheRS. 
     
     
         9 . The transgenic plant of  claim 8 , wherein the  E. Coli  PheRS-α is encoded by a polynucleotide having the nucleic acid sequence set forth in SEQ ID NO:1 and the  E. Coli  PheRS-β is encoded by a polynucleotide having the nucleic acid sequence set forth in SEQ ID NO:2. 
     
     
         10 . The transgenic plant of  claim 8 , wherein the  E. Coli  PheRS-α comprises the amino acid sequence set forth in SEQ ID NO:3; the  E. Coli  PheRS-β comprises the amino acid sequence set forth in SEQ ID NO:4; the  T. thermophilus  PheRS-α comprises the amino acid sequence set forth in SEQ ID NO:5 and the  T. thermophilus  PheRS-β comprises the amino acid sequence set forth in SEQ ID NO:6. 
     
     
         11 . The transgenic plant of  claim 1 , wherein the polynucleotide encoding the aaRS or fragment thereof further comprises a nucleic acid sequence encoding a targeting peptide selected from the group consisting of a mitochondrial targeting peptide and a chloroplast targeting peptide. 
     
     
         12 . The transgenic plant of  claim 11 , wherein said plant comprises a combination of (a) the polynucleotide encoding the aaRS or a fragment thereof further comprising the nucleic acid sequence encoding a mitochondrial targeting peptide and (b) the polynucleotide encoding the aaRS or a fragment thereof further comprising the nucleic acid sequence encoding a chloroplast targeting peptide. 
     
     
         13 . A plant seed produced by the transgenic plant of  claim 1 , wherein the seed is used for breeding a transgenic plant comprising at least one exogenous polynucleotide encoding an aminoacyl tRNA synthase (aaRS) or a fragment thereof having an editing module capable of hydrolyzing tRNA misacylated with non-protein amino acid analog, wherein the transgenic plant is resistant to the non-protein amino acid analog and salts thereof. 
     
     
         14 . A tissue culture comprising at least one transgenic cell of the plant of  claim 1  or a protoplast derived therefrom, wherein the tissue culture regenerates a transgenic plant comprising at least one exogenous polynucleotide encoding an aminoacyl tRNA synthase (aaRS) or a fragment thereof having an editing module capable of hydrolyzing tRNA misacylated with non-protein amino acid analog, the transgenic plant is resistant to the non-protein amino acid analog and salts thereof. 
     
     
         15 . A method for producing a transgenic plant resistant to phytotoxic non-protein amino acid analog or a salt thereof, the method comprising the steps of:
 (a) transforming a plant cell with at least one exogenous polynucleotide encoding an aminoacyl tRNA synthetase (aaRS) or a fragment thereof the aaRS or fragment thereof comprising an editing module capable of hydrolyzing non-protein aminoacylated tRNA; and   (b) regenerating the transformed cell into a transgenic plant resistant to the phytotoxic non-protein amino acid analog or a salt thereof.   
     
     
         16 . The method of  claim 15 , wherein the non-protein amino acid analog is meta-tyrosine (m-Tyr) compound and the aaRS is phenylalanyl-tRNA synthetase (PheRS). 
     
     
         17 . The method of  claim 16 , wherein the PheRS is a heterotetrameric bacterial PheRS composed of two PheRS-α and two PheRS-β strands. 
     
     
         18 . The method of  claim 17 , wherein the bacterial PheRS is  Escherichia coli  ( E. coli ) PheRS or  Thermus thermophilus  PheRS. 
     
     
         19 . The method of  claim 18 , wherein the  E. Coli  PheRS-α is encoded by a polynucleotide having the nucleic acid sequence set forth in SEQ ID NO:1 and wherein the  E. Coli  PheRS-β is encoded by a polynucleotide having the nucleic acid sequence set forth in SEQ ID NO:2. 
     
     
         20 . The method of  claim 18 , wherein the  E. Coli  PheRS-α comprises the amino acid sequence set forth in SEQ ID NO:3; the  E. Coli  PheRS-β comprises the amino acid sequence set forth in SEQ ID NO:4; the  T. thermophilus  PheRS-α comprises the amino acid sequence set forth in SEQ ID NO:5 and the  T. thermophilus  PheRS-β comprises the amino acid sequence set forth in SEQ ID NO:6. 
     
     
         21 . The method of  claim 15 , wherein the polynucleotide encoding the aaRS or fragment thereof further comprises a nucleic acid sequence encoding a targeting peptide selected from the group consisting of a mitochondrial targeting peptide and a chloroplast targeting peptide. 
     
     
         22 . The method of  claim 21 , wherein the plant cell is transformed with a combination of (a) the polynucleotide encoding the aaRS or a fragment thereof further comprising the nucleic acid sequence encoding a mitochondrial targeting peptide and (b) the polynucleotide encoding the (aaRS) or a fragment thereof further comprising the nucleic acid sequence encoding a chloroplast targeting peptide. 
     
     
         23 . A transgenic plant produced by the method of  claim 15 , wherein the plant is resistant to the phytotoxic non-protein amino acid.