Insect resistance using inhibition of gene expression
First Claim
1. A chimeric gene comprising the following operably linked DNA:
- (a) a plant-expressible promoter;
(b) a DNA region which when transcribed yields a double-stranded RNA molecule capable of reducing the expression of an essential gene of a plant sap-sucking insect, said RNA molecule comprising a first and second RNA region wherein;
(i) said first RNA region comprises a nucleotide sequence of at least 19 consecutive nucleotides having at least about 94% sequence identity to the nucleotide sequence of said endogenous gene;
(ii) said second RNA region comprises a nucleotide sequence complementary to said at least 19 consecutive nucleotides of said first RNA region;
(iii) said first and second RNA region are capable of base-pairing to form a double stranded RNA molecule between at least said 19 consecutive nucleotides of said first and second region;
(c) optionally, a 3′
end region comprising transcription termination and polyadenylation signals functioning in cells of said plant.
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Accused Products
Abstract
The current invention provides methods to silence insect genes by using unpackaged dsRNA or siRNA, in one embodiment such dsRNA or siRNA is present in plant vascular tissue, preferably phloem, more particularly phloem sap, and the insect is a plant sap-sucking insect. Also provided are DNA sequences which when transcribed yield a double-stranded RNA molecule capable of reducing the expression of an essential gene of a plant sap-sucking insect, methods of using such DNA sequences and plants or plant cells transformed with such DNA sequences. Also provided is the use of cationic oligopeptides that facilitate the entry of dsRNA or siRNA molecules in insect cells, such as plant sap-sucking insect cells.
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Citations
28 Claims
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1. A chimeric gene comprising the following operably linked DNA:
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(a) a plant-expressible promoter;
(b) a DNA region which when transcribed yields a double-stranded RNA molecule capable of reducing the expression of an essential gene of a plant sap-sucking insect, said RNA molecule comprising a first and second RNA region wherein;
(i) said first RNA region comprises a nucleotide sequence of at least 19 consecutive nucleotides having at least about 94% sequence identity to the nucleotide sequence of said endogenous gene;
(ii) said second RNA region comprises a nucleotide sequence complementary to said at least 19 consecutive nucleotides of said first RNA region;
(iii) said first and second RNA region are capable of base-pairing to form a double stranded RNA molecule between at least said 19 consecutive nucleotides of said first and second region;
(c) optionally, a 3′
end region comprising transcription termination and polyadenylation signals functioning in cells of said plant. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 17, 25, 26, 28)
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- 10. A method to silence a gene of a plant sap-sucking insect, comprising applying to the feed of said plant sap-sucking insect unpackaged, naked dsRNA or siRNA which is targeted to an essential plant sap-sucking gene.
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13. A method to silence a gene in an plant sap-sucking insect, comprising:
- adding naked, unpackaged dsRNA or siRNA to the diet or feed of said plant sap-sucking insect, wherein said dsRNA or siRNA targets said gene.
- 14. A method of controlling plant sap-sucking insects, comprising expressing in the phloem of a plant dsRNA that targets an essential plant sap-sucking insect gene.
- 18. A method of identifying gene function in a plant sap-sucking insect, comprising the step of applying naked, unpackaged dsRNA targeting a plant sap-sucking insect gene to the diet of said insect, and evaluating phenotypic or biochemical changes in said insect.
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19. A method of identification of novel targets for insecticidal compounds, comprising the steps of:
- a) applying naked, unpackaged dsRNA or siRNA molecules to the feed or diet of a plant-sap sucking insect;
b) analyzing which genes when silenced confer lethality to said insect, c) cloning and characterizing said genes thus analyzed;
d) identifying compounds that disrupt or inactivate said gene or the RNA or protein encoded thereby; and
e) contacting said compounds with said insect or feed or diet of said insect to confirm the pesticidal nature of said compound. - View Dependent Claims (27)
- a) applying naked, unpackaged dsRNA or siRNA molecules to the feed or diet of a plant-sap sucking insect;
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20. Phloem of a plant, comprising siRNA targeted to an aphid essential gene.
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21. Phloem sap of a plant, comprising siRNA targeted to an aphid essential gene.
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22. An aphid gene comprising the sequence of any one of SEQ ID NO:
- 5 to 8, SEQ ID NO;
11 or SEQ ID NO;
12.
- 5 to 8, SEQ ID NO;
Specification