DNA encoding a plant lipase, transgenic plants and a method for controlling senescence in plants

• US 6,774,284 B1
• Filed: 07/05/2000
• Issued: 08/10/2004
• Est. Priority Date: 06/26/1998
• Status: Expired due to Fees

• Current Assignee: Thomson Licensing (Vantiva SA)
• Original Assignee: Senesco, Inc.
• Inventors: Hudak, Katalin, Thompson, John E., Wang, Tzann-Wei, Hong, Yuwen
• Primary Examiner(s): MEHTA, ASHWIN D
• Application Number:
  US09/610,104
• Time in Patent Office: 1,497 Days
• Field of Search: 536/23.6, 435/69.1, 435/320.1, 435/410, 435/419, 435/468, 435/411, 800/278, 800/281, 800/283, 800/285, 800/286, 800/287, 800/290, 800/295, 800/298, 800/317.4, 800/323.3
• US Class Current: 800/290
• CPC Class Codes: C12N 15/8249 involving ethylene biosynthesis, senescence or fruit development, e.g. modified tomato ripening, cut flower shelf-life
• CPC Class Codes: C12N 15/8266 Abscission; Dehiscence; Senescence
• CPC Class Codes: C12N 15/8273 for drought, cold, salt resistance
• CPC Class Codes: C12N 9/18 Carboxylic ester hydrolases (3.1.1)
• CPC Class Codes: C12N 9/20 Triglyceride splitting, e.g. by means of lipase
• CPC Class Codes: C12Q 1/6895 for plants, fungi or algae
• CPC Class Codes: C12Q 2600/158 Expression markers
• CPC Class Codes: C40B 30/04 by measuring the ability to specifically bind a target molecule, e.g. antibody-antigen binding, receptor-ligand binding
• CPC Class Codes: C40B 40/08 Libraries containing RNA or DNA which encodes proteins, e.g. gene libraries

Abstract

Regulation of expression of senescence in plants is achieved by integration of a gene or gene fragment encoding senescence-induced lipase into the plant genome in antisense orientation. The carnation and Arabidopsis genes encoding senescence-induced lipase are identified and the nucleotide sequences are used to modify senescence in transgenic plants.

Claims

1. An isolated DNA molecule encoding senescence-induced lipase, wherein the DNA molecule has at least 90% sequence similarity to SEQ ID NO:

1, or SEQ ID NO;

18, and wherein the DNA molecule encodes an amino acid sequence that has senescence-induced lipase activity of SEQ ID NO;

2 or SEQ ID NO;

19.

3. The isolated DNA molecule of claim 1 wherein the isolated DNA molecule comprises a nucleotide sequence encoding the amino acid sequence set forth in SEQ ID NO:

4.

2. An isolated DNA molecule encoding senescence-induced lipase wherein said DNA molecule comprises SEQ ID NO:

1.

4. An isolated DNA molecule encoding senescence-induced lipase wherein said DNA molecule comprises SEQ ID NO:

18.

5. A vector for transformation of plant cells comprising

(a) an antisense polynucleotide complementary to SEQ ID NO:

1, or SEQ D NO;

18, and

(b) regulatory sequences operatively linked to the antisense polynucleotide such that the antisense polynucleotide is expressed in a plant cell into which it is transformed.

6. The vector according to claim 5 wherein the regulatory sequences comprise a promoter and a transcription termination region.

7. The vector according to claim 5 wherein the regulatory sequences comprise a constitutive promoter.

8. The vector according to claim 5 wherein the regulatory sequences comprise a plant tissue-specific promoter.

9. The vector according to claim 5 wherein the regulatory sequences comprise a senescence-induced plant promoter.

10. The vector according to claim 5 wherein the regulatory sequences comprise a viral promoter.

34. A transgenic plant cell comprising the vector according to claim 5.

38. A transgenic plant comprising the vector according to claim 5.

51. Progeny of the plant of claim 38, wherein said progeny comprise said vector.

11. An isolated antisense polynucleotide encoding an RNA molecule, wherein said polynucleotide has at least 90% sequence similarity to the complement of SEQ ID NO:

1, or SEQ ID NO;

18, wherein the polynucleotide inhibits expression of an endogenous senescence-induced lipase gene when expressed in a plant cell.

12. The antisense polynucleotide according to claim 11, wherein the antisense polynucleotide is the complement of the nucleotide sequence set forth in SEQ ID NO:

1.

13. The antisense polynucleotide according to claim 11, wherein the antisense polynucleotide is the complement of the nucleotide sequence set forth in SEQ ID NO:

18.

14. The antisense polynucleotide according to claim 11, wherein the plant cell is a carnation plant cell.

15. The antisense polynucleotide according to claim 11, wherein the plant cell is an Arabidopsis plant cell.

16. The antisense polynucleotide according to claim 11, wherein the plant cell is a tomato plant cell.

36. A plasmid comprising a replication system functional in a prokaryotic host and the antisense polynucleotide according to claim 11.

37. A plasmid comprising a replication system functional in Agrobacterium and the antisense polynucleotide according to claim 11.

17. A vector comprising

(a) a DNA molecule encoding senescence-induced lipase, wherein the DNA molecule has at least 90% sequence similarity to SEQ ID NO:

1 or SEQ ID NO;

18; and

(b) regulatory sequences operatively linked to the DNA molecule,

(c) wherein the DNA molecule encodes an amino acid sequence that has senescence-induced lipase activity of SEQ ID NO;

2 or SEQ ID NO;

19.

18. A bacterial cell transformed with the vector according to claim 17.

35. A transgenic plant cell comprising the vector according to claim 17.

19. A method for inhibiting expression of an endogenous senescence-induced lipase in a plant, said method comprising

(1) integrating into a plant genome a vector comprising

(A) an antisense polynucleotide having at least 90% sequence similarity to the complement of SEQ ID NO:

1 or SEQ ID NO;

18; and

(B) regulatory sequences operatively linked to the antisense polynucleotide such that the antisense polynucleotide is expressed; and

(2) growing said plant, whereby the antisense polynucleotide is transcribed and binds to an RNA transcript of an endogenous senescence-induced lipase gene, whereby expression of the senescence-induced lipase gene is inhibited.

20. The method according to claim 19 wherein said inhibition results in delayed senescence of the plant.

21. The method according to claim 19 wherein said inhibition results in increased resistance of said plant to environmental stress-induced senescence.

22. The method according to claim 19 wherein said inhibition results in enhanced biomass of said plant.

23. The method according to claim 19 wherein said inhibition results in increased seed yield in said plant.

24. The method according to claim 19 wherein the regulatory sequences comprise a constitutive promoter active in the plant.

25. The method according to claim 19 wherein the regulatory sequences comprise a double 35S promoter.

26. The method according to claim 19 wherein the regulatory sequences comprise a tissue specific promoter active in the plant.

27. The method according to claim 19 wherein the regulatory sequences comprise a senescence-induced promoter active in the plant.

28. The method according to claim 19 wherein said plant is selected from the group consisting of fruit bearing plants, flowering plants, vegetables, agronomic crop plants and forest species.

29. The method according to claim 19 wherein the plant is a tomato.

30. The method according to claim 19 wherein the plant is a carnation.

31. A method for inhibiting the expression of an endogenous senescence-induced lipase gene or genes in a plant cell, said method comprising

(1) integrating into a genome of at least one cell of the plant a vector comprising

(A) an isolated DNA molecule encoding an exogenous senescence-induced lipase, wherein the DNA molecule comprises SEQ ID NO:

1 or SEQ ID NO;

18, or wherein the DNA molecule has at least 90% sequence similarity to SEQ ID NO;

1 or SEQ ID NO;

18, and wherein the DNA molecule encodes an amino acid sequence that has senescene-induced lipase activity of SEQ ID NO;

2 or SEQ ID NO;

19; and

(B) regulatory sequences operatively linked to the DNA molecule such that the DNA molecule is transcribed, and

(2) growing said plant, wherein the endogenous senescence-induced lipase is co-suppressed.

32. The method according to claim 31 wherein the regulatory sequences comprise a constitutive promoter.

33. A method of delaying age-related senescence and environmental stress-related senescence in a plant, said method comprising

(1) integrating a vector into a plant genome, the vector comprising

(A) an antisense polynucleotide having at least 90% sequence similarity to the complement of SEQ ID NO:

1 or SEQ ID NO;

18, and

(B) regulatory sequences operatively linked to the antisense polynucleotide such that the antisense polynucleotide is expressed; and

(2) growing said plant, whereby the antisense polynucleotide is transcribed and binds to an RNA transcript of an endogenous senescence-induced lipase gene, whereby expression of the senescence-induced lipase gene is inhibited,

(3) whereby age-related and environment stress-related senescence is delayed.

39. A plant and progeny thereof, wherein the plant is derived from a cell having inhibited or reduced expression of senescence-induced lipase, wherein said cell is produced by

(1) integrating into a cell genome a vector comprising

(A) an antisense polynucleotide having at least 90% sequence similarity to the complement of SEQ ID NO:

1 or SEQ ID NO;

18; and

wherein the polynucleotide inhibits expression of an endogenous senescence-induced lipase gene, and

(B) regulatory sequences operatively linked to the antisense polynucleotide such that the antisense polynucleotide is expressed; and

(2) growing said plant, whereby said antisense polynucleotide is transcribed and binds an RNA transcript of an endogenous senescence induced lipase gene whereby expression of said senescence-induced lipase gene is inhibited; and

(3) wherein the plant and progeny thereof comprise the vector.

40. The plant or progeny thereof according to claim 39 wherein the plant or progeny thereof is a tomato plant.

41. The plant or progeny thereof according to claim 39 wherein the plant or progeny thereof is a carnation plant.

42. A method of inhibiting seed aging, said method comprising

(1) integrating into a plant genome a vector comprising

(A) an antisense polynucleotide having at least 90% sequence similarity to the complement SEQ ID NO:

1 or SEQ ID NO;

18; and

(B) regulatory sequences operatively linked to the antisense polynucleotide; and

(2) growing said plant, whereby said antisense polynucleotide is transcribed and binds to an RNA transcript of an endogenous senescence-induced lipase gene whereby expression of said senescence-induced lipase gene is inhibited,

(3) whereby seed aging is inhibited.

43. A method of increasing seed yield from a plant, said method comprising

(1) integrating into a plant genome a vector comprising

(A) an antisense polynucleotide having at least 90% sequence similarity to the complement of SEQ ID NO:

1 or SEQ ID NO;

18; and

(B) regulatory sequences operatively linked to the antisense polynucleotide; and

(2) growing said plant, whereby said antisense polynucleotide is transcribed and binds to an RNA transcript of an endogenous senescence-induced lipase gene whereby expression of said senescence-induced lipase gene is inhibited,

(3) whereby seed yield is increased.

44. A vector comprising

(a) a DNA molecule encoding a senescence-induced lipase, wherein the DNA molecule comprises SEQ ID NO:

1 or SEQ ID NO;

18; and

(b) regulatory sequences operatively linked to the DNA molecule.

45. A method for inhibiting expression of an endogenous senescence-induced lipase in a plant, said method comprising

(1) integrating into a plant genome a vector comprising

(A) an antisense polynucleotide complementary to a DNA molecule encoding the endogenous senescence-induced lipase, wherein the DNA molecule encoding the endogenous senescence-induced lipase comprises SEQ ID NO:

1 or SEQ ID NO;

18; and

(B) regulatory sequences operatively linked to the antisense polynucleotide such that the antisense polynucleotide is expressed; and

(2) growing said plant, whereby the antisense polynucleotide is transcribed and binds to an RNA transcript of said DNA molecule, whereby expression of the DNA molecule is inhibited.

46. A method for inhibiting the expression of an endogenous senescence-induced lipase gene or genes in a plant, said method comprising

(1) integrating into a genome of at least one cell of the plant a vector comprising

(A) an isolated DNA molecule encoding exogenous senescence-induced lipase comprising SEQ ID NO:

1 or SEQ ID NO;

18; and

(B) regulatory sequences operatively linked to the DNA molecule such that the DNA molecule is transcribed, and

(2) growing said plant, wherein the endogenous senescence-induced lipase is co-suppressed.

47. A method of altering age-related senescence and environmental stress-related senescence in a plant, said method comprising

(1) integrating a vector into a plant genome, the vector comprising

(A) an antisense polynucleotide complementary to SEQ ID NO:

1 or SEQ ID NO;

18; and

(B) regulatory sequences operatively linked to the antisense polynucleotide such that the antisense polynucleotide is expressed; and

(2) growing said plant, whereby the antisense polynucleotide is transcribed and binds to an RNA transcript of an endogenous senescence-induced lipase gene, whereby expression of the senescence-induced lipase gene is inhibited.

48. A plant or progeny thereof, wherein the plant is regenerated from a cell having inhibited or reduced expression of senescence-induced lipase, wherein said cell is produced by

(1) integrating into the cell genome a vector comprising

(A) an antisense polynucleotide complementary to SEQ ID NO:

1 or SEQ ID NO;

18; and

(B) regulatory sequences operatively linked to the antisense polynucleotide such that the antisense polynucleotide is expressed; and

(2) growing said plant, whereby said antisense polynucleotide is transcribed and binds an RNA transcript of an endogenous senescence induced lipase gene whereby expression of said senescence-induced lipase gene is inhibited; and

(3) wherein the plant or progeny thereof comprise the vector.

49. A method of inhibiting seed aging, said method comprising

(1) integrating into a plant genome a vector comprising

(A) an antisense polynucleotide complementary to SEQ ID NO:

1 or SEQ ID NO;

18; and

(B) regulatory sequences operatively linked to the antisense polynucleotide; and

(2) growing said plant, whereby said antisense polynucleotide is transcribed and binds to an RNA transcript of an endogenous senescence-induced lipase gene whereby expression of said senescence-induced lipase gene is inhibited,

(3) whereby seed aging is inhibited.

50. A method of increasing seed yield from a plant, said method comprising

(1) integrating into a plant genome a vector comprising

(A) an antisense polynucleotide complementary to SEQ ID NO:

1 or SEQ ID NO;

18; and

(B) regulatory sequences operatively linked to the antisense polynucleotide; and

(2) growing said plant, whereby said antisense polynucleotide is transcribed and binds to an RNA transcript of an endogenous senescence-induced lipase gene whereby expression of said senescence-induced lipase gene is inhibited,

(3) whereby seed yield is increased.