METHODS AND COMPOSITIONS FOR CONFERRING AND/OR ENHANCING HERBICIDE TOLERANCE USING PROTOPORPHYRINOGEN OXIDASE OR VARIANT THEREOF

• US 20190330651A1
• Filed: 06/15/2017
• Published: 10/31/2019
• Est. Priority Date: 06/16/2016
• Status: Active Grant

• Current Assignee: FarmHannong Co., Ltd.
• Original Assignee: FarmHannong Co., Ltd.
• Inventors: SUNG, Soon-Kee, YOON, Joonseon, HAN, Yunjung, AHN, Young Ock, PARK, Joonghyuk, HONG, Myoung-ki
• Granted Patent: US 10,844,395 B2
• Time in Patent Office: Days
• CPC Class Codes: C07K 14/195 from bacteria
• CPC Class Codes: C12N 1/12 Unicellular algae; Culture media therefor as new plants A01H13/00
• CPC Class Codes: C12N 15/8274 for herbicide resistance
• CPC Class Codes: C12N 15/8275 Glyphosate
• CPC Class Codes: C12N 15/8277 Phosphinotricin
• CPC Class Codes: C12N 9/001 acting on the CH-CH group of donors (1.3)
• CPC Class Codes: C12Y 103/03004 Protoporphyrinogen oxidase (1.3.3.4)

Abstract

Provided is technology for conferring enhanced herbicide tolerance and/or enhancing herbicide tolerance of plants and/or algae using a protoporphyrinogen oxidase derived from prokaryotes or its amino acid variants.

Claims

1. A polypeptide selected from the followings:

a polypeptide comprising an amino acid sequence wherein at least one selected from the group consisting of N59, S60, R89, F161, V165, A167, Q184, P303, V305, F324, L327, I340, F360, and I408 of the amino acid sequence of SEQ ID NO;

2 is independently deleted or substituted with an amino acid which is selected from the group consisting of M(Met), V(Val), I(Ile), T(Thr), L(Leu), C(Cys), A(Ala), S(Ser), F(Phe), P(Pro), W(Trp), N(Asn), Q(Gln), G(Gly), Y(Tyr), D(Asp), E(Glu), R(Arg), H(His), and K(Lys), and is different from the original amino acid at the corresponding position;

a polypeptide comprising an amino acid sequence wherein at least one selected from the group consisting of R101, F171, V175, A177, G194, P316, V318, F337, L340, I353, and F373 of the amino acid sequence of SEQ ID NO;

4 is independently deleted or substituted with an amino acid which is selected from the group consisting of M(Met), V(Val), I(Ile), T(Thr), L(Leu), C(Cys), A(Ala), S(Ser), F(Phe), P(Pro), W(Trp), N(Asn), Q(Gln), G(Gly), Y(Tyr), D(Asp), E(Glu), R(Arg), H(His), and K(Lys), and is different from the amino acid at the corresponding position in the wild type; and

a polypeptide consisting of an amino acid sequence having 95% or higher homology with the amino acid sequence of said polypeptides.

2. The polypeptide of claim 1, wherein the polypeptide is selected from the group consisting of,

a polypeptide comprising of an amino acid sequence wherein at least one selected from the group consisting of N59, S60, R89, F161, V165, A167, Q184, P303, V305, F324, L327, I340, F360, and I408 of the amino acid sequence of SEQ ID NO:

2 is independently substituted with an amino acid which is selected from the group consisting of M(Met), V(Val), I(Ile), T(Thr), L(Leu), C(Cys), A(Ala), S(Ser), R(Arg), W(Trp), and G(Gly);

a polypeptide comprising an amino acid sequence wherein at least one selected from the group consisting of R101, F171, V175, A177, G194, P316, V318, F337, L340, I353, and F373 of the amino acid sequence of SEQ ID NO;

4 is independently substituted with an amino acid which is selected from the group consisting of M(Met), V(Val), I(Ile), T(Thr), L(Leu), C(Cys), A(Ala), E(Glu), Q(Gln), K(Lys), R(Arg), H(His), and N(Asn); and

a polypeptide consisting of an amino acid sequence having 95% or higher homology with the amino acid sequence of said polypeptides.

3. The polypeptide of claim 1, wherein the polypeptide is selected from the group consisting of,

a polypeptide comprising an amino acid sequence comprising at least one amino acid mutation selected from the group consisting of F360M, F360V, F360I, F360T, F360L, F360C, A167C, A167L, A167I, P303L, V305L, V305M, V305T, N59T, S60T, R89A, R89L, R89V, F161A, V165S, V165C, Q184G, F324V, L327T, I340T, I408R, and I408W in the amino acid sequence of SEQ ID NO:

2;

a polypeptide comprising an amino acid sequence comprising at least one amino acid mutation selected from the group consisting of F373M, F373V, F373I, F373T, F373L, F373C, F373N, F373H, A177C, A177L, A177I, P316A, P316L, V318L, V318M, R101A, F171A, V175C, V175L, G194E, G194Q, G194M, G194K, G194R, F337V, L340T, and I353T in the amino acid sequence of SEQ ID NO;

4; and

a polypeptide consisting of an amino acid sequence having 95% or higher homology with the amino acid sequence of said polypeptides.

4. The polypeptide of claim 3, wherein the polypeptide is selected from the group consisting of,

a polypeptide comprising an amino acid sequence comprising amino acid mutation of F360M, F360V, F360I, F360T, F360L, F360C, A167C, A167L, P303L, N59T, 560T, R89A, R89L, R89V, F161A, V165S, V165C, A167I, Q184G, V305L, V305M, V305T, F324V, L327T, I340T, I408R, I408W, P303L+V305L, N59T+F360V, S60T+V165S+F360M, S60T+V165S+F360I, S60T+I340T+F360I, R89A+F360M, R89A+F360I, R89A+F360L, R89L+F360I, R89V+F360I, R89A+A167L+F360M, R89A+V305T+F360M, V165S+F360M, V165S+F360I, V165S+F360L, V165S+F360V, V165C+F360M, V165C+A167C+F360M, V165C+A167I+F360M, V165C+A167L+F360M, A167L+F360M, A167L+F360I, A167C+F360M, A167C+F360I, A167I+F360M, V305M+F360M, V305T+F360I, V305L+F360M, I408R+F360M, or I408W+F360M in the amino acid sequence of SEQ ID NO:

2;

a polypeptide comprising an amino acid sequence comprising amino acid mutation of F373M, F373V, F373I, F373T, F373L, F373C, F373N, F373H, A177C, A177L, A177I, P316A, P316L, V318L, V318M, R101A, F171A, V175C, V175L, G194E, G194Q, G194M, G194K, G194R, F337V, L340T, I353T, P316L+V318L, P316A+V318L, R101A+F373M, A177C+F373M, A177I+F373M, A177L+F373M, A177L+F373I, A177L+F373L, A177L+F373T, A177L+F373V, A177C+F373T, A177C+F373V, V175L+F373M, G194E+F373M, G194Q+F373M, G194M+F373M, G194K+F373M, G194R+F373M, or V318M+F373M in the amino acid sequence of SEQ ID NO;

4; and

a polypeptide consisting of an amino acid sequence having 95% or higher homology with the amino acid sequence of said polypeptides.

5. A polynucleotide encoding the polypeptide of claim 1.

6. A recombinant vector comprising the polynucleotide of claim 5.

7. A recombinant cell comprising the recombinant vector of claim 6.

8. A composition for conferring or enhancing herbicide tolerance of a plant or algae, comprising at least one selected from the group consisting of the polypeptide of SEQ ID NO:

2;

the polypeptide of SEQ ID NO;

4;

the polypeptide of claim 1;

a polynucleotide encoding the polypeptide;

a recombinant vector comprising the polynucleotide; and

a recombinant cell comprising the recombinant vector.

9. The composition of claim 8, wherein the herbicide is an herbicide inhibiting protoporphyrinogen oxidase.

10. The composition of claim 9, wherein the herbicide is at least one selected from the group consisting of pyrimidinediones, diphenyl-ethers, phenylpyrazoles, N-phenylphthalimides, phenylesters, thiadiazoles, oxadiazoles, triazolinones, oxazolidinediones, pyraclonil, flufenpyr-ethyl and profluazol.

11. The composition of claim 10, wherein the herbicide is at least one selected from the group consisting of butafenacil, saflufenacil, benzfendizone, tiafenacil, fomesafen, oxyfluorfen, aclonifen, acifluorfen, bifenox, ethoxyfen, lactofen, chlomethoxyfen, chlorintrofen, fluoroglycofen-ethyl, halosafen, pyraflufen-ethyl, fluazolate, flumioxazin, cinidon-ethyl, flumiclorac-pentyl, fluthiacet, thidiazimin, oxadiargyl, oxadiazon, carfentrazone, sulfentrazone, azafenidin, pentoxazone, pyraclonil, flufenpyr-ethyl, profluazol, phenopylate, carbamate analogues of phenopylate, and agriculturally acceptable salt thereof.

12. The composition of claim 8, wherein the plant or algae further comprise a second herbicide-tolerant polypeptide or a gene encoding thereof, and tolerance to the second herbicide is conferred or enhanced.

13. The composition of claim 12, wherein the second herbicide is selected from the group consisting of glyphosate, glufosinate, dicamba, 2,4-D (2,4-dichlorophenoxyacetic acid), isoxaflutole, ALS(acetolactate synthase)-inhibiting herbicide, photosystem II-inhibiting herbicide, phenylurea-based herbicide, bromoxynil-based herbicide, and combinations thereof.

14. The composition of claim 12, wherein the second herbicide is at least one selected from the group consisting of,

glyphosate herbicide-tolerant EPSPS (glyphosate tolerant 5-enolpyruvylshikimate-3-phosphate synthase), GOX (glyphosate oxidase), GAT (glyphosate-N-acetyltransferase) or glyphosate decarboxylase;

glufosinate herbicide-tolerant PAT (phosphinothricin-N-acetyltransferase);

dicamba herbicide-tolerant DMO (Dicamba monooxygenase);

2,4-D (2,4-dichlorophenoxyacetic acid) herbicide-tolerant 2,4-D monooxygenase or AAD (aryloxyalkanoate Dioxygenase);

ALS (acetolactate synthase)-inhibiting sulfonylurea-based herbicide-tolerant ALS (acetolactate synthase), AHAS (acetohydroxyacid synthase) or AtAHASL Atahasl (acetohydroxyacid synthase large subunit);

photosystem II-inhibiting herbicide-tolerant photosystem II protein D1;

phenylurea herbicide-tolerant Cytochrome P450;

plastid-inhibiting herbicide-tolerant HPPD (Hydorxylphenylpyruvate dioxygenase);

bromoxynil herbicide-tolerant Nitrilase; and

combinations thereof.

15. The composition of claim 12, wherein the gene encoding the second herbicide-tolerant polypeptide is at least one selected from the group consisting of,

glyphosate herbicide-tolerant cp4 epsps, epsps (AG), mepsps, 2mepsps, goxv247, gat4601 or gat4621 gene;

glufosinate herbicide-tolerant bar or pat gene;

dicamba herbicide-tolerant dmo gene;

2,4-D (2,4-Dichlorophenoxyacetic acid) herbicide-tolerant AAD-1 or AAD-12 gene;

isoxaflutole herbicide-tolerant HPPDPF W336 gene;

sulfonylurea herbicide-tolerant ALS, Csr1, Csr1-1, Csr1-2, GM-HRA, S4-HRA, Zm-HRA, SurA or SurB gene;

photosystem II-inhibiting herbicide-tolerant psbA gene;

phenylurea herbicide-tolerant CYP76B1 gene;

bromoxynil herbicide-tolerant bxn gene; and

combinations thereof.

16. A transformant of a plant or algae having herbicide tolerance, or a clone or progeny thereof, comprising the polypeptide of SEQ ID NO:

2, the polypeptide of SEQ ID NO;

4, the polypeptide of claim 1, or a polynucleotide encoding thereof.

17. The transformant, clone or progeny thereof of claim 16, wherein the transformant is plant cell, protoplast, callus, hypocotyl, seed, cotyledon, shoot, or whole plant.

18. A method of preparing plants or algae having herbicide tolerance, the method comprising transforming algae, or plant cell, protoplast, callus, hypocotyl, seed, cotyledon, shoot, or whole plant, with the polypeptide of SEQ ID NO:

2, the polypeptide of SEQ ID NO;

4, the polypeptide of claim 1, or a polynucleotide encoding thereof.

19. A method of conferring or enhancing herbicide tolerance of plants or algae, the method comprising transforming algae, or plant cell, protoplast, callus, hypocotyl, seed, cotyledon, shoot, or whole plant, with the polypeptide of SEQ ID NO:

2, the polypeptide of SEQ ID NO;

4, the polypeptide of claim 1, or a polynucleotide encoding thereof.

20. A method of controlling weeds in a cropland, the method comprising, providing the cropland with a plant comprising the polypeptide of SEQ ID NO:

2, the polypeptide of SEQ ID NO;

4, the polypeptide of claim 1, or a polynucleotide encoding thereof, and

applying an effective dosage of protoporphyrinogen oxidase-inhibiting herbicide to the cropland.

21. The method of claim 20, wherein the step of applying an effective dosage of protoporphyrinogen oxidase-inhibiting herbicide to the cropland is performed by applying an effective dosage of two or more kinds of protoporphyrinogen oxidase-inhibiting herbicides sequentially or simultaneously.

22. The method of claim 20,

wherein the plant further comprises the second herbicide-tolerant polypeptide or a gene encoding thereof, and

the step of applying an effective dosage of protoporphyrinogen oxidase-inhibiting herbicide to the cropland is performed by applying an effective dosage of the protoporphyrinogen oxidase-inhibiting herbicide and a second herbicide are applied sequentially or simultaneously.

23. A method of removing an undesired aquatic organism from a culture medium, the method comprising,

providing a culture medium with algae comprising the polypeptide of SEQ ID NO:

2, the polypeptide of SEQ ID NO;

4, the polypeptide of claim 1 or a polynucleotide encoding thereof, and

applying an effective dosage of protoporphyrinogen oxidase-inhibiting herbicide to the culture medium.