Hypervirulent mutant of
View Patent ↗This disclosure relates to a mutant Agrobacterium tumefaciens that is functionally deleted for the atu1060 gene that codes for the cyclic di-GMP synthase Atu1060, as well as methods for its use in transforming plants with desired transgenes. Such bacteria are more virulent than currently used strains of A. tumefaciens , and thus can be used to transform a wider variety of plants, such as plants that are traditionally recalcitrant to such transformation.
1. An isolated Agrobacterium tumefaciens ( A. tumefaciens ) bacterium, wherein a cyclic di-GMP synthase atu1060 gene is functionally deleted.
2. The isolated A. tumefaciens bacterium of claim 1 , wherein the bacterium is a recombinant bacterium comprising an exogenous nucleic acid molecule inserted into a cyclic di-GMP synthase atu1060 gene of the bacterium, thereby functionally deleting the cyclic di-GMP synthase atu1060 gene.
3. The isolated bacterium of claim 1 , wherein the bacterium is A. tumefaciens strain NTL4, NT1, GV2260, GV3101, GV3122, GV3166, GV3850, ACH5, EHA101, EHA105, or C58.
4. The isolated bacterium of claim 1 , wherein the isolated bacterium induces tumors on a host plant at a minimum infection dose at least 5-fold lower than those required by the same strain of A. tumefaciens bacterium having a functional cyclic di-GMP synthase atu1060 gene.
5. The isolated bacterium of claim 1 , wherein the isolated bacterium attaches to glass slides with at least 4-fold greater efficiency than the same strain of A. tumefaciens bacterium having a functional cyclic di-GMP synthase atu1060 gene.
6. The isolated bacterium of claim 1 , wherein the isolated bacterium demonstrates polar lectin binding with at least 4-fold greater efficiency than the same strain of A. tumefaciens bacterium having a functional cyclic di-GMP synthase atu1060 gene.
7. The isolated bacterium of claim 1 , wherein the functional deletion is achieved by removal of at least a portion of the atu1060 gene.
8. The isolated bacterium of claim 1 , wherein the atu1060 gene in a wild-type A. tumefaciens bacterium encodes a protein having at least 95% sequence identity to the amino acid sequence shown in SEQ ID NO: 1.
9. The isolated bacterium of claim 1 , wherein the atu1060 gene in a wild-type A. tumefaciens bacterium comprises the nucleic acid sequence shown in SEQ ID NO: 2.
10. The isolated bacterium of claim 1 , wherein the bacterium does not comprise a nucleic acid sequence comprising SEQ ID NO: 2.
11. The isolated bacterium of claim 1 , wherein the bacterium does not comprise an amino acid sequence having at least 95% sequence identity to the amino acid sequence shown in SEQ ID NO: 1.
12. The isolated bacterium of claim 1 , further comprising a transgene that provides a benefit to a plant.
13. The isolated bacterium of claim 12 , wherein the transgene is a gene that confers resistance to a pest, drought, or herbicide.
14. A method of transforming a plant, comprising:
contacting a plant cell with the isolated bacterium of claim 1 , under conditions that permit the bacterium to infect the plant cell, thereby transforming the plant.
15. The method of claim 14 , wherein the plant cell is a corn cell, wheat cell, rice cell, soybean cell, alfalfa cell, tomato cell, cotton cell, or canola cell.
16. The isolated bacterium of claim 1 , wherein the bacterium is an A. tumefaciens biovar 1 isolate.
17. The isolated bacterium of claim 16 , wherein the bacterium is A. tumefaciens strain C58.
18. The isolated bacterium of claim 1 , wherein the atu1060 gene in a wild-type A. tumefaciens bacterium encodes a protein having at least 92% sequence identity to the amino acid sequence shown in SEQ ID NO: 1.
19. The isolated bacterium of claim 1 , wherein the bacterium does not comprise an amino acid sequence having at least 92% sequence identity to the amino acid sequence shown in SEQ ID NO: 1.
20. The isolated bacterium of claim 1 , wherein the atu1060 gene in a wild-type A. tumefaciens bacterium comprises a nucleic acid sequence having at least 90% sequence identity to the nucleic acid sequence shown in SEQ ID NO: 2.
21. The isolated bacterium of claim 1 , wherein the bacterium does not comprise a nucleic acid sequence having at least 90% sequence identity to the nucleic acid sequence shown in SEQ ID NO: 2.