Expression constructs, vird2 mutant agrobacterium strains, and methods of use thereof
AssigneePurdue Research Foundation
InventorsStanton Gelvin, Lan-Ying Lee
Life of this patent
Estimating the term…
Abstract
Agrobacterium strains with mutations in the VirD2 gene and protein and expression constructs and genome-editing systems for leveraging the same. Methods are also provided for Agrobacterium-mediated incorporation of exogenous expressible nucleic acids into a host plant material using the Agrobacterium strains, expression constructs, and/or genome-editing systems.
Claims55
An Agrobacterium strain comprising a mutant VirD2 gene that effects transient transformation of a plant, the mutant VirD2 strain encoding a wild-type VirD2 protein except for:
- (i) a mutation at a threonine at position 319 of the VirD2 protein;
- (ii) a mutation at a threonine at position 319 of the VirD2 protein, a mutation at a histidine at position 402 of the VirD2 protein, and an insertion of 30 to 40 amino acid residues at a terminal end of the VirD2 protein resulting from a truncation mutation at a glutamine at position 425;
- or
- (iii) an insertion of 1 to 20 amino acid residues after position 326 of the VirD2 protein, which affects a downstream frameshift.
An expression construct comprising a mutant VirD2 gene that encodes a wild-type Agrobacterium VirD2 protein except for:
- (i) a mutation at a threonine at position 319 of the VirD2 protein;
- (ii) a mutation at a threonine at position 319 of the VirD2 protein, a mutation at a histidine at position 402 of the VirD2 protein, and an insertion of 30 to 40 amino acid residues at a terminal end of the VirD2 protein due to a truncation mutation at a glutamine at position 425;
- or
- (iii) an insertion of 1 to 22 amino acid residues after position 326 of the VirD2 protein, which affects a downstream frameshift.
A genome editing system comprising:
- an expression construct comprising a mutant Agrobacterium VirD2 gene that encodes an Agrobacterium wild-type VirD2 protein except for:
- (i) a mutation at a threonine at position 319 of the VirD2 protein,
- (ii) a mutation at a threonine at position 319 of the VirD2 protein, a mutation at a histidine at position 402 of the VirD2 protein, and an insertion of 30 to 40 amino acid residues at a terminal end of the VirD2 protein resulting from a truncation of a glutamine at position 425 of the VirD2 protein, or
- (iii) an insertion of 1 to 20 amino acid residues after position 326 of the VirD2 protein, which affects a downstream frameshift;
- and
- a sequence-specific nuclease, a DNA polymerase, and/or a DNA polymerase recruitment protein, or an expression construct comprising a nucleotide sequence encoding the sequence-specific nuclease, the DNA polymerase and/or the DNA polymerase recruitment protein.
Dependencies are inferred from the claim text.