US2019316128A1PendingUtilityA1

DELIVERY OF dsRNA TO ARTHROPODS

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Assignee: COMMW SCIENT IND RES ORGPriority: Jul 6, 2001Filed: Jun 10, 2019Published: Oct 17, 2019
Est. expiryJul 6, 2021(expired)· nominal 20-yr term from priority
Inventors:Steven WhyardFiona Helen CameronMinoo MoghaddamTrevor J. Lockett
A61P 33/00A61P 31/04A61P 31/12A61P 31/00A61P 43/00A61K 38/00C12Y 302/01031C12N 2310/14C12N 2320/50C12N 2320/32C12N 15/113A61K 31/713C12N 15/1137C12N 15/111A01N 57/16A01K 2217/05A01K 67/0337Y02A40/146A01K 67/65
69
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Claims

Abstract

The invention is to methods of gene silencing in arthropods using dsRNA. The method is include contacting the arthropod with, and/or directly feeding the arthropod, the dsRNA to the arthropods to deliver the dsRNA to arthropod tissues. It is envisaged that the methods of the invention will have use in determining the biological function of genes in arthropods. Methods of pest control of arthropods, and of protecting arthropods against parasites and predators are provided. Transgenic arthropods expressing dsRNA molecules are also provided by the present invention.

Claims

exact text as granted — not AI-modified
1 . A method of determining the biological function of a target RNA in an arthropod comprising delivering to the arthropod a dsRNA molecule which specifically reduces the level of the target RNA and/or the production of a protein encoded by the target RNA in a cell of the arthropod, and assessing the effect of the dsRNA on at least one biological function of the arthropod. 
     
     
         2 . The method of  claim 1 , wherein the dsRNA is delivered by a process comprising contacting the arthropod with the dsRNA. 
     
     
         3 . The method of  claim 2 , wherein said contacting comprises wholly or partially soaking the arthropod in a composition comprising the dsRNA. 
     
     
         4 . The method of  claim 1 , wherein the dsRNA is delivered by a process comprising feeding the dsRNA to the arthropod. 
     
     
         5 . The method according to any one of  claims 1  to  4 , wherein the dsRNA is delivered in a composition comprising a transfection promoting agent. 
     
     
         6 . The method of  claim 5 , wherein the transfection promoting agent is a lipid-containing compound. 
     
     
         7 . The method of  claim 6 , wherein the lipid-containing compound is selected from the group consisting of; Lipofectamine, Cellfectin, DMRIE-C, DOTAP and Lipofectin. 
     
     
         8 . The method of  claim 6 , wherein the lipid-containing compound is a Tris cationic lipid. 
     
     
         9 . The method of  claim 8 , wherein the Tris cationic lipid is selected from the group consisting of: CS096, CS102, CS129, CS078, CS051, CS027, CS041, CS042, CS060, CS039, and CS015. 
     
     
         10 . The method according to any one of  claims 5  to  9 , wherein the composition further comprises a nucleic acid condensing agent. 
     
     
         11 . The method of  claim 10 , wherein the nucleic acid condensing agent is selected from the group consisting of spermidine and protamine sulfate. 
     
     
         12 . The method of  claim 1 , wherein the dsRNA is delivered by a process comprising feeding a transgenic organism expressing the dsRNA to the arthropod. 
     
     
         13 . The method of  claim 12 , wherein the transgenic organism is a transgenic plant. 
     
     
         14 . The method of  claim 1 , wherein the dsRNA is delivered by a process comprising contacting the arthropod with a virus expressing the dsRNA. 
     
     
         15 . The method according to any one of  claims 1  to  14 , wherein the dsRNA comprises a nucleotide sequence having at least 90% identity to at least a portion of the sequence of the target RNA. 
     
     
         16 . The method according to any one of  claims 1  to  14 , wherein the dsRNA comprises a nucleotide sequence having at least 99% identity to at least a portion of the sequence of the target RNA. 
     
     
         17 . The method according to any one of  claims 1  to  16 , wherein the arthropod is an insect. 
     
     
         18 . A composition comprising dsRNA and a transfection promoting agent, wherein said dsRNA comprises a nucleotide sequence that it is at least 90% identical to the sequence of a target RNA, wherein the target RNA is selected from the group consisting oft a naturally-occurring arthropod RNA, a naturally-occurring RNA of an organism that is a pathogen carried by an arthropod, a naturally-occurring RNA of a virus that infects an arthropod, an RNA copy of a naturally-occurring DNA virus that infects an arthropod, and a naturally-occurring RNA of a bacterium that infects an arthropod. 
     
     
         19 . The composition of  claim 18 , wherein the transfection promoting agent is a lipid-containing compound. 
     
     
         20 . The composition of  claim 19 , wherein the lipid-containing compound is selected from the group consisting of; Lipofectamine, Cellfectin, DMRIE-C, DOTAP and Lipofectin. 
     
     
         21 . The composition of  claim 19 , wherein the lipid-containing compound is a Tris cationic lipid. 
     
     
         22 . The composition of  claim 21 , wherein the Tris cationic lipid is selected from the group consisting of: CS096, CS102, CS129, CS078, CS051, CS027, CS041, CS042, CS060, CS039, and C8015. 
     
     
         23 . The composition according to any one of  claims 18  to  22 , wherein the composition further comprises a nucleic acid condensing agent. 
     
     
         24 . The composition of  claim 23 , wherein the nucleic acid condensing agent is selected from the group consisting of: spermidine and protamine sulfate. 
     
     
         25 . The composition according to any one of  claims 18  to  24 , wherein the composition is formulated such that it can be applied to an area inhabited by a population of arthropods. 
     
     
         26 . The composition according to any one of  claims 18  to  25 , wherein the composition further comprises an agriculturally acceptable carrier. 
     
     
         27 . The composition according to any one of  claims 18  to  26 , wherein the composition is formulated as a bait. 
     
     
         28 . A method of controlling an arthropod pest comprising delivering to the arthropod dsRNA by a process comprising contacting the arthropod with said dsRNA or feeding said dsRNA to the arthropod, for a time and under conditions sufficient for said dsRNA, or a degradation product thereof, to specifically reduce the level of a target RNA and/or the production of a protein encoded by the target RNA in a cell of the arthropod, wherein the target RNA or the protein is important for arthropod survival, development and/or reproduction. 
     
     
         29 . The method of  claim 28 , wherein the dsRNA is delivered in a composition according to any one of  claims 18  to  27 . 
     
     
         30 . A method for controlling a pathogen transmitted by an arthropod, the method comprising delivering to the arthropod dsRNA by a process comprising contacting the arthropod with said dsRNA or feeding said dsRNA to said arthropod, for a time and under conditions sufficient for said dsRNA, or a degradation product thereof, to specifically reduce the level of a target RNA and/or the production of a protein encoded by the target RNA in a cell of the pathogen, wherein the target RNA or the protein is important for pathogen survival, development and/or reproduction. 
     
     
         31 . The method of  claim 30 , wherein the dsRNA is delivered in a composition according to any one of  claims 18  to  27 . 
     
     
         32 . The method of  claim 30  or  claim 31 , wherein the pathogen is selected from the group consisting of: fungi, protozoans, bacteria and viruses. 
     
     
         33 . A method of protecting an arthropod against a pathogen, parasite or predatory organism, the method comprising delivering to the arthropod dsRNA by a process comprising contacting the arthropod with said dsRNA or feeding said dsRNA to said arthropod, for a time and under conditions sufficient for said dsRNA, or a degradation product thereof, to so specifically reduce the level of a target RNA and/or the production of a protein encoded by the target RNA in a cell of the pathogen, parasite or predatory organism, wherein the target RNA or the protein is important for the survival, development and/or reproduction of the pathogen, parasite or predatory organism. 
     
     
         34 . The method of  claim 33 , wherein the dsRNA is delivered in a composition according to any one of  claims 18  to  27 . 
     
     
         35 . A transgenic organism comprising a heterologous nucleic acid(s) which is transcribed to produce a dsRNA, wherein the portion of the dsRNA which is double stranded is about 21 to about 50 base pairs in length. 
     
     
         36 . The transgenic organism of  claim 35 , wherein the dsRNA comprises a nucleotide sequence having at least 90% identity to at least a portion of the sequence of a target RNA selected from the group consisting of a naturally-occurring arthropod RNA, a naturally-occurring RNA of an organism that is a pathogen carried by an arthropod, a naturally-occurring RNA of a virus that infects an arthropod, an RNA copy of a naturally-occurring DNA virus that infects an arthropod, and a naturally-occurring RNA of a bacterium that infects an arthropod. 
     
     
         37 . The transgenic organism of  claim 35  or  claim 36 , wherein the portion of the dsRNA which is double stranded is about 21 to about 23 base pairs in length. 
     
     
         38 . The transgenic organism according to any one of  claims 35  to  37 , wherein the organism is selected from the group consisting of: plants and arthropods.

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