Polynucleotides and methods for making plants resistant to fungal pathogens

Abstract

This invention relates to polynucleotide sequences encoding a gene that can confer resistance to the plant pathogen Colletotrichum, which causes anthracnose stalk rot, leaf blight and top dieback in corn and other cereals. It further relates to plants and seeds of plants carrying chimeric genes comprising said polynucleotide sequences, which enhance or confer resistance to the plant pathogen Colletotrichum, and processes of making said plants and seeds. The invention further presents sequences that can be used as molecular markers that in turn can be used to identify the region of interest in corn lines resulting from new crosses and to quickly and efficiently introgress the gene from corn lines carrying said gene into other corn lines that do not carry said gene, in order to make them resistant to Colletotrichum and resistant to stalk rot.

1 . An isolated polynucleotide comprising a polynucleotide selected from the group consisting of: 
 (a) a nucleotide sequence encoding a polypeptide conferring or enhancing resistance to  Colletotrichum,  wherein the polypeptide has an amino acid sequence of at least 75% identity, when compared to SEQ ID NO:3 based on the Needleman-Wunsch alignment algorithm;    (b) a complement of the nucleotide sequence, wherein the complement and the nucleotide sequence consist of the same number of nucleotides and are 100% complementary; and    (c) a nucleotide sequence encoding a polypeptide conferring or enhancing resistance to  Colletotrichum,  wherein the polypeptide has an amino acid sequence of at least 75% identity, when compared to the sequence of the plasmid deposited as Patent Deposit No. NRRL B-30895 based on the Needleman-Wunsch alignment algorithm.    
     
     
         2 . The polynucleotide of  claim 1 , wherein the amino acid sequence of the polypeptide and the amino acid sequence of SEQ ID NO: 3 have at least 90% identity based on the Needleman-Wunsch alignment algorithm.  
     
     
         3 . The polynucleotide of  claim 1 , wherein the amino acid sequence of the polypeptide and the amino acid sequence of SEQ ID NO: 3 have at least 95% identity based on the Needleman-Wunsch alignment algorithm.  
     
     
         4 . The polynucleotide of  claim 1 , wherein the nucleotide sequence comprises SEQ ID NO: 3.  
     
     
         5 . The polynucleotide of  claim 1 , wherein the nucleotide sequence comprises the sequence of the plasmid deposited as Patent Deposit No. NRRL B-30895.  
     
     
         6 . A vector comprising the polynucleotide of  claim 1 .  
     
     
         7 . A recombinant DNA construct comprising the polynucleotide of  claim 1  operably linked to at least one regulatory sequence.  
     
     
         8 . A process for transforming a host cell, comprising transforming the host cell with the polynucleotide of  claim 1 .  
     
     
         9 . The process of  claim 8  wherein the host cell is a plant cell.  
     
     
         10 . The process of  claim 9  wherein the plant cell is from maize.  
     
     
         11 . A plant cell comprising the recombinant DNA construct of  claim 7 .  
     
     
         12 . A plant cell comprising the polynucleotide of  claim 1 .  
     
     
         13 . The plant cell of  claim 12 , wherein said plant cell is from maize.  
     
     
         14 . A process for producing a plant comprising transforming a plant cell with the recombinant DNA construct of  claim 7  and regenerating a plant from the transformed plant cell.  
     
     
         15 . A plant comprising the recombinant DNA construct of  claim 7 .  
     
     
         16 . The plant of  claim 15 , wherein said plant is maize.  
     
     
         17 . A seed comprising the recombinant DNA construct of  claim 7 .  
     
     
         18 . The seed of  claim 17 , wherein said seed is maize seed.  
     
     
         19 . A process of conferring or enhancing resistance to  Colletotrichum,  comprising transforming a plant with the recombinant DNA construct of  claim 7 , thereby conferring or enhancing resistance to  Colletotrichum.    
     
     
         20 . A process of determining the presence or absence of the polynucleotide of  claim 1  in a corn plant, comprising at least one of: 
 (a) isolating nucleic acid molecules from said corn plant and amplifying sequences homologous to the polynucleotide of  claim 1 , or    (b) isolating nucleic acid molecules from said corn plants and performing a Southern hybridization, or    (c) isolating proteins from said corn plant and performing a western blot using antibodies to the Rcg1 protein, or    (d) isolating proteins from said corn plant and performing an ELISA assay using antibodies to the Rcg1 protein, or    (e) demonstrating the presence of mRNA sequences derived from the Rcg1 mRNA transcript and unique to Rcg1, thereby determining the presence of the polynucleotide of  claim 1  in said corn plant.    
     
     
         21 . A process of determining the presence or absence of the Rcg1 locus in a corn plant, comprising at least one of: 
 (a) isolating nucleic acid molecules from said corn plant and amplifying sequences homologous to the polynucleotide of  claim 1 , or    (b) isolating nucleic acid molecules from said corn plant and performing a Southern hybridization, or    (c) isolating proteins from said corn plant and performing a western blot using antibodies to the Rcg1 protein, or    (d) isolating proteins from said corn plant and performing an ELISA assay using antibodies to the Rcg1 protein, or    (e) demonstrating the presence of mRNA sequences derived from the Rcg1 mRNA transcript and unique to Rcg1, thereby determining the presence of the Rcg1 locus in said corn plant.    
     
     
         22 . A process of altering the level of expression of a protein capable of conferring resistance to  Colletotrichum  in a plant cell comprising: 
 (a) transforming a plant cell with the recombinant DNA construct of  claim 7;  and    (b) growing the transformed plant cell under conditions that are suitable for expression of the recombinant DNA construct wherein expression of the recombinant DNA construct results in production of altered levels of a protein capable of conferring resistance to  Colletotrichum  in the transformed plant cell.    
     
     
         23 . A process of altering the level of expression of a protein capable of conferring resistance to stalk rot in a plant cell comprising: 
 (a) transforming a plant cell with the recombinant DNA construct of  claim 7;  and    (b) growing the transformed plant cell under conditions that are suitable for expression of the recombinant DNA construct wherein expression of the recombinant DNA construct results in production of altered levels of a protein capable of conferring resistance to stalk rot in the transformed plant cell.    
     
     
         24 . A process of altering the level of expression of a protein capable of conferring resistance to  Colletotrichum  in a plant comprising: 
 (a) transforming a plant cell with the recombinant DNA construct of  claim 7;  and    (b) regenerating a transformed plant from the transformed plant cell; and    (c) growing the transformed plant under conditions that are suitable for expression of the recombinant DNA construct wherein expression of the recombinant DNA construct results in production of altered levels of a protein capable of conferring resistance to  Colletotrichum  in the transformed plant.    
     
     
         25 . A process of altering the level of expression of a protein capable of conferring resistance to stalk rot in a plant comprising: 
 (a) transforming a plant cell with the recombinant DNA construct of  claim 7;  and    (b) regenerating the transformed plant from the transformed plant cell; and    (c) growing the transformed plant under conditions that are suitable for expression of the recombinant DNA construct wherein expression of the recombinant DNA construct results in production of altered levels of a protein capable of conferring resistance to stalk rot in the transformed plant.