Mutated hydroxyphenylpyruvate dioxygenase, DNA sequence and isolation of plants which contain such a gene and which are tolerant to herbicides

• US 6,245,968 B1
• Filed: 02/18/1999
• Issued: 06/12/2001
• Est. Priority Date: 11/07/1997
• Status: Expired due to Term

• Current Assignee: Bayer SAS (Bayer AG)
• Original Assignee: Aventis Cropscience SA
• Inventors: Rodgers, Matthew, Sailland, Alain, Dumas, Florence, Bourdon, Hélène, Boudec, Philippe
• Primary Examiner(s): Benzion, Gary
• Assistant Examiner(s): MEHTA, ASHWIN D
• Application Number:
  US09/252,292
• Time in Patent Office: 845 Days
• Field of Search: 435/69.1, 435/320.1, 435/410, 435/418, 435/419, 435/468, 435/325, 435/471, 435/483, 435/484, 435/488, 435/69.7, 536/23.6, 536/23.7, 536/23.1, 800/21, 800/260, 800/278, 800/295, 800/298, 800/300
• US Class Current: 800/278
• CPC Class Codes: C12N 15/8274 for herbicide resistance
• CPC Class Codes: C12N 9/0069 acting on single donors with incorporation of molecular oxygen, i.e. oxygenases (1.13)

Abstract

The present invention relates to a nucleic acid sequence encoding a mutated hydroxyphenylpyruvatedioxygenase (HPPD) which exhibits improved tolerance to HPPD inhibitors, to a chimeric gene which comprises this sequence as the coding sequence, and to its use for isolating or obtaining plants which are resistant to certain herbicides.

Claims

1. An isolated nucleic acid which encodes an enzymatically-active, mutated HPPD, which HPPD has been mutated in its C-terminal part to be less sensitive to HPPD inhibitors than the native, unmutated HPPD.

2. A nucleic acid according to claim 1, wherein a linking peptide is located between 5 and 15 amino acids upstream of Asp161, with reference to Pseudomonas HPPD (SEQ ID NO:

31).

3. A nucleic acid according to claim 2, wherein the C-terminal part consists of the part between the linking peptide and the C-terminal end of the HPPD.

4. A nucleic acid according to claim 1, wherein mutation is an amino acid substitution.

5. The nucleic acid according to claim 4, wherein the substitution is effected on at least one amino acid of the C-terminal part which is common to several HPPD sequences.

6. The nucleic acid of claim 4, comprising one or more substitutions selected from the group consisting of Pro215Leu, Gly336Glu, Gly336Trp, Gly336Ile and Ala340Gly.

7. The nucleic acid of claim 4, wherein the substitution adds amino acids which exhibit greater steric hindrance than the amino acids being replaced.

8. The nucleic acid of claim 4, wherein the substituted amino acids are replaced with glutamine (Gln), glutamic acid (Glu), leucine (Leu), isoleucine (Ile) or tryptophan (Trp).

9. The nucleic acid according to claim 1, wherein the mutation is carried out on the C-terminal part between the positions corresponding to positions 290 and 350 of the Pseudomonas sequence (SEQ ID NO:

31).

10. The nucleic acid according to claim 9, wherein the mutation is a substitution of an amino acid at one of positions 298, 332, 333, 334, 336 or 340.

11. The nucleic acid of claim 9, wherein the mutation is an amino acid substitution at Pro215, Gly298, Gly332, Phe333, Gly334, Gly336 or Ala340.

12. The nucleic acid of claim 11, wherein the mutation is at Pro215, Gly336 or Ala340.

13. A nucleic acid of claim 1, wherein the HPPD contains, in its C-terminal part, the following peptide sequence:

14. A nucleic acid sequence of claim 13, wherein at least one of Xaa and Xab represents Leu, Glu, Trp or Ile.

15. A nucleic acid sequence of claim 14, wherein Xab represents Glu, Trp or Ile.

16. A nucleic acid sequence of claim 15, wherein Xab represents Trp.

17. A chimeric gene which comprises the nucleic acid of claim 1 and regulatory elements allowing its expression in a host, wherein the host is plant, a plant cell, or a bacteria.

18. The chimeric gene of claim 17, wherein the host is plant.

19. The chimeric gene of claim 18, further comprising sequence encoding a transit peptide.

20. A plant cell which contains the chimeric gene of claim 19.

21. A plant transformation vector comprising the chimeric gene of claim 17.

22. A plant cell which contains the chimeric gene of claim 18.

23. A plant which contains the chimeric gene of claim 19, which is tolerant to HPPD herbicides.

24. A seed of a plant as claimed in claim 23.

25. A plant which contains the chimeric gene of claim 17, which is tolerant to HPPD herbicides.

26. A seed of a plant as claimed in claim 25.

27. A method which comprises growing the plant of claim 25 in a field, and applying an HPPD herbicide to the field.

28. A method which comprises growing the plant of claim 23 in a field, and applying an HPPD herbicide to the field.

29. A plant which contains the nucleic acid of claim 1 and which is tolerant to HPPD herbicides.