dsRNA as insect control agent

• US 9,528,123 B2
• Filed: 08/27/2014
• Issued: 12/27/2016
• Est. Priority Date: 01/12/2006
• Status: Active Grant

• Current Assignee: deVGen NV (China National Chemical Corporation)
• Original Assignee: deVGen NV (China National Chemical Corporation)
• Inventors: Raemaekers, Romaan, Kubler, Laurent, Vanbleu, Els, Bogaert, Thierry Andre Olivier Eddy
• Primary Examiner(s): IBRAHIM, MEDINA AHMED
• Application Number:
  US14/470,868
• Publication Number:
  US 20140373197A1
• Time in Patent Office: 853 Days
• Field of Search: None
• US Class Current: 1/1
• CPC Class Codes: A01N 57/16 containing heterocyclic radicals
• CPC Class Codes: A01N 61/00 Biocides, pest repellants or attractants, or plant growth regulators containing substances of unknown or undetermined composition, e.g. substances characterised only by the mode of action
• CPC Class Codes: C07K 14/43527 from ticks
• CPC Class Codes: C07K 14/4354 from nematodes
• CPC Class Codes: C12N 15/113 Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing when used in plants C12N15/8218
• CPC Class Codes: C12N 15/8282 for fungal resistance
• CPC Class Codes: C12N 15/8285 for nematode resistance
• CPC Class Codes: C12N 15/8286 for insect resistance
• CPC Class Codes: C12N 2310/14 interfering N.A.
• CPC Class Codes: Y02A 40/146 Genetically Modified [GMO] plants, e.g. transgenic plants

Abstract

The present invention relates to methods for controlling pest infestation using double stranded RNA molecules. The invention provides methods for making transgenic plants that express the double stranded RNA molecules, as well as pesticidal agents and commodity products produced by the inventive plants.

Claims

1. An isolated double stranded RNA molecule comprising annealed complementary strands, wherein at least one of said strands comprises a polyribonucleotide selected from the group consisting of:

(i) polyribonucleotides complementary to at least 50 contiguous nucleotides of the target gene of SEQ ID NO;

3,

(ii) polyribonucleotides complementary to at least 50 contiguous nucleotides of a target gene encoding the amino acid sequence of SEQ ID NO;

4, and

(iii) polyribonucleotides having at least 90% sequence identity with the polyribonucleotides of (i) or (ii),

wherein said double stranded RNA molecule has pesticidal activity.

2. A polynucleotide or set of polynucleotides encoding a double stranded RNA molecule according to claim 1.

3. The double stranded RNA molecule of claim 1, wherein ingestion of said double stranded RNA molecule by a plant insect pest inhibits the growth of said insect pest.

4. The double stranded RNA molecule of claim 1, wherein ingestion of said double stranded RNA molecule inhibits expression of a nucleotide sequence substantially complementary to said polyribonucleotide.

5. A cell transformed with a polynucleotide encoding a double stranded RNA molecule according to claim 1.

6. The cell of claim 5, wherein said cell is a plant cell.

7. A plant transformed with a polynucleotide encoding a double stranded RNA molecule according to claim 1.

8. The plant of claim 7, wherein said double stranded RNA molecule inhibits a pest biological activity.

9. The plant of claim 7, wherein said double stranded RNA molecule inhibits expression of a target gene.

10. The plant of claim 9 wherein said target gene is an insect, nematode or fungal gene.

11. The plant of claim 7, wherein said plant is cytoplasmic male sterile.

12. The plant of claim 7, wherein said plant further comprises or expresses a pesticidal agent selected from the group consisting of a patatin, a Bacillus thuringiensis insecticidal protein, a Xenorhabdus insecticidal protein, a Photorhabdus insecticidal protein, a Bacillus laterosporous insecticidal protein, and a Bacillus sphearicus insecticidal protein.

13. The plant of claim 12 wherein said Bacillus thuringiensis insecticidal protein is selected from the group consisting of a Cry1, a Cry3, a TIC851, a CryET170, a Cry22, a binary insecticidal protein CryET33 and CryET34, a binary insecticidal protein CryET80 and CryET76, a binary insecticidal protein TIC100 and TIC101, and a binary insecticidal protein PS149B1.

14. The plant of claim 7, wherein said plant selected from the group consisting of alfalfa, apple, apricot, artichoke, asparagus, avocado, banana, barley, beans, beet, blackberry, blueberry, broccoli, Brussels sprouts, cabbage, canola, carrot, cassava, cauliflower, a cereal, celery, cherry, citrus, clementine, coffee, corn, cotton, cucumber, eggplant, endive, eucalyptus, figs, grape, grapefruit, groundnuts, ground cherry, kiwifruit, lettuce, leek, lemon, lime, pine, maize, mango, melon, millet, mushroom, nut, oat, okra, onion, orange, an ornamental plant or flower or tree, papaya, parsley, pea, peach, peanut, peat, pepper, persimmon, pineapple, plantain, plum, pomegranate, potato, pumpkin, radicchio, radish, rapeseed, raspberry, rice, rye, sorghum, soy, soybean, spinach, strawberry, sugar beet, sugarcane, sunflower, sweet potato, tangerine, tea, tobacco, tomato, a vine, watermelon, wheat, yams and zucchini.

15. The plant of claim 6, wherein said plant is resistant against infestation by an insect selected from the group consisting of Leptinotarsa spp., Lema spp., Epitrix spp., Epicauta spp., Epilachna spp., Phaedon spp., Nilaparvata spp., Laodelphax spp., Nephotettix spp., Sogatella spp., Acheta spp., Blissus spp., Scotinophora spp., Acrosternum spp., Parnara spp., Chilo spp., Chilotraea spp., Sesamia spp., Tryporyza spp., Cnaphalocrocis spp., Agromyza spp., Diatraea spp., Narnaga spp., Xanthodes spp., Spodoptera spp., Mythimna spp., Helicoverpa spp., Colaspis spp., Lissorhoptrus spp., Echinocnemus spp., Diclodispa spp., Oulema spp., Sitophilus spp., Pachydiplosis spp., Hydrellia spp., Chlorops spp., Diabrotica spp., Ostrinia spp., Agrotis spp., Elasmopalpus spp., Melanotos spp., Cyclocephala spp., Popillia spp., Chaetocnema spp., Sphenophorus spp., Rhopalosiphum spp., Anuraphis spp., Melanoplus spp., Hylemya spp., Anaphothrips spp., Solenopsis spp., Helicoverpa spp., Pectinophora spp., lianas spp., Heliothis spp., Anthonomus spp., Pseudatomoscelis spp., Trialeurodes spp., Bemisia spp., Aphis spp., Lygus spp., Euschistus spp., Chlorochroa spp., Nezara spp., Thrips spp., Frankliniella spp., Empoasca spp., Myzus spp., Paratrioza spp., Conoderus spp., Phthorimaea spp., Macrosiphum spp., Thyanta spp., Phthorimaea spp., Helicoverpa spp., Keiferia spp., Limonius spp., Manduca spp., Liriomyza spp., Drosophilia spp., Carabus spp., Chironomus spp., Ctenocephalides spp., Diaprepes spp., Ips spp., Tribolium spp., Glossina spp., Anopheles spp., Helicoverpa spp., Acyrthosiphon spp., Apis spp., Homalodisca spp., Aedes spp., Bombyx spp., Locusta spp., Boophilus spp., Acanthoscurria spp., Diploptera spp., Heliconius spp., Curculio spp., Plutella spp., Amblyomma spp., Anteraea spp., and Armigeres spp..

16. A seed, part, tissue, or cell of the plant of claim 7, wherein said seed, part, tissue or cell comprises a double stranded RNA molecule according to claim 1.

17. A product produced from the plant of claim 7, wherein said product comprises said double stranded RNA molecule.

18. The product of claim 17, wherein said product is selected from the group consisting of food, feed, fiber, paper, meal, protein, starch, flour, silage, coffee, tea, and oil.

19. A pesticide comprising the product of claim 17, said product expressing said double stranded RNA molecule.

20. A method for controlling or preventing insect growth comprising providing an insect pest with a product of a plant transformed with a polynucleotide encoding the double stranded RNA molecule of claim 1.

21. A method for producing a plant resistant against a plant pathogenic organism comprising:

a) transforming a plant cell with a polynucleotide encoding the double stranded RNA molecule of claim 1,

b) regenerating a plant from the transformed plant cell; and

c) growing the transformed plant under conditions suitable for the expression of said double stranded RNA molecule,

wherein said grown transformed plant is resistant to said plant pathogenic organism compared to an untransformed plant.

22. A method for improving yield, comprising:

a) transforming a plant cell with a polynucleotide encoding the double stranded RNA molecule of claim 1,

b) regenerating a plant from the transformed plant cell; and

c) growing the transformed plant under conditions suitable for the expression of said double stranded RNA molecule from said polynucleotide,

wherein said expression inhibits feeding by a plant pathogenic organism and reduces loss of yield due to pest infestation.

23. A transgenic plant resistant to an insect pest comprising the double stranded RNA molecule of claim 1.

24. The transgenic plant according to claim 23 further comprising or expressing a pesticidal agent selected from the group consisting of a patatin, a Bacillus thuringiensis insecticidal protein, a Xenorhabdus insecticidal protein, a Photorhabdus insecticidal protein, a Bacillus laterosporous insecticidal protein, and a Bacillus sphearicus insecticidal protein.

25. The transgenic plant of claim 24 wherein said Bacillus thuringiensis insecticidal protein is selected from the group consisting of a Cry1, a Cry3, a TIC851, a CryET170, a Cry22, a binary insecticidal protein CryET33 and CryET34, a binary insecticidal protein CryET80 and CryET76, a binary insecticidal protein TIC100 and TIC101, and a binary insecticidal protein PS149B1.

26. A pesticide comprising a plant of claim 7 or a seed, part, tissue, or cell thereof, wherein said plant, seed, part, tissue or cell thereof expresses said double stranded RNA molecule.

27. A method for controlling or preventing insect growth comprising providing an insect pest with a plant transformed with a polynucleotide encoding the double stranded RNA molecule of claim 1.

28. The method according to claim 27, wherein the insect pest is selected from the group consisting of Leptinotarsa spp., Tribolium spp., Myzus spp., Nilaparvata spp., Chilo spp., and Acheta spp.

29. The method according to claim 28, wherein the insect pest is selected from the group consisting of Leptinotarsa decemlineata, Tribolium castaneum, Myzus persicae, Nilaparvata lugens, Chilo suppressalis, and Acheta domesticus.

30. The plant of claim 15, wherein said plant is resistant against infestation by an insect selected from the group consisting

of Leptinotarsa decemlineata (Colorado potato beetle), Leptinotarsa juncta (false potato beetle), Leptinotarsa texana (Texan false potato beetle), Lema trilineata (three-lined potato beetle), Epitrix cucumeris (potato flea beetle), Epitrix hirtipennis (flea beetle), Epitrix tuberis (tuber flea beetle), Epicauta vittata (striped blister beetle),

Epilachna varivetis (mexican bean beetle), Phaedon cochieariae (mustard leaf beetle), Nitaparvata lugens (brownplanthopper), Laodelphax striatellus (small brown planthopper), Nephoteltix virescens, Nephotettix cincticeps (green leafhopper), Nephotettik nigropictus (rice leafhopper), Sogatella furcifera (white-backed planthopper), Acheta domesticus (house cricket), Blissus leucopterus leucopterus (chinch bug), Scotinophora vermidulate (rice blackbug), Acrosternum hilare (green stink bug), Parnara guttata (rice skipper), Chilo suppressalis (rice striped stem borer), Chilo auricilius (gold-fringed stem borer), Chilo polychrysus (dark-headed stem borer), Chilotraea polychrysa (rice stalk borer), Sesamia inferens (pink rice borer), Tryporyza innotata (white rice borer), Tryporyza incertulas (yellow rice borer), Cnaphalocrocis medinalis (rice leafroller), Agromyza oryzae (leafminer), Agromyza parvicornis (corn blot leafminer), Diatraea saccharalis (sugarcane borer), Diatraea grandiosella (southwestern corn borer), Narnaga aenescens (green rice caterpillar), Xanthodes transversa (green caterpillar), Spodoptera frugiperda (fall armyworm), Spodoptera exigua (beet armyworm), Spodoptera littoralis (climbing cutworm), Spodoptera praefica (western yellowstriped armyworm), Helicoverpa zea (corn earworm), Colaspis brunnea (grape colaspis), Lissorhoptrus oryzophilus (rice water weevil), Echinocnemus squamos (rice plant weevil), Diciodispa armigera (rice hispa), Oulema oryzae (leaf beetle), Sitophilus oryzae (rice weevil), Pachydiplosis oryzae (rice gall midge), Hydrellia griseola (small rice leafminer), and Hydrellia sasakil (rice stem maggot), Chlorops oryzae (stem maggot), Diabrotica virgifera virgifera (western corn rootworm), Diabrotica barberi (northern corn rootworm), Diabrotica undecimpunctala howardi (southern corn rootworm), Diabrotica virgifera zeae (Mexican corn rootworm);

Diabrotica balteata (banded cucumber beetle), Ostrinia nubilalis (European corn borer), Agrotis ipsilon (black cutworm), Elasmopalpus lignosellus (lesser cornstalk borer), Melanotus wireworms, Cyclocephaia borealis (northern masked chafer), Cyclocephala immaculata (southern masked chafer), Popillia japonica (Japanese beetle), Chaetocnema pulicaria (corn flea beetle), Sphenophorus maidis (maize billbua), Rhopalosiphum maidis (corn leaf aphid), Anuraphis maidiradicis (corn root aphid), Melanoplus femurrubrum (redlegged grasshopper), Melanoplus differentialis (differential grasshopper), Melanoplus sanguinipes (migratory grasshopper), Hylemya platura (seedcorn maggot), Anaphothrips obscrurus (grass thrips), Solenopsis milesta (thief ant), Helicoverpa zea (cotton bollworm), Helicoverpa armigera (American bollworm)), Pectinophora gossypiella (pink bollworm), Heliothis virescens (tobacco budworm), Anthonomus grandis (boll weevil), Pseudatomoscelis seriatus (cotton fleahopper), Trialeurodes abutiloneus (banded-winged whitefly), Trialeurodes vaporariorum (greenhouse whitefly), Bemisia argentifoln (silverleaf whitefly), Aphis gossypii (cotton aphid), Lygus lineolaris (tarnished plant bug), Lygus hesperus (western tarnished plant bug), Euschistus conspersus (consperse stink bug), Chlorochroa sayi (Say stinkbug), Nezara viridula (southern green stinkbug), Thrips tabaci (onion thrips), Frankliniella fusca (tobacco thrips), Frankliniella occidentalis (western flower thrips), Empoasca fabae (potato leafhopper), Myzus persicae (green peach aphid), Paratrioza cockerelli, Conoderus falli (southern potato wireworm), and C. vespertinus (tobacco wireworm), Phthorimaea operculella (potato tuberworm), Macrosiphum euphorbiae (potato aphid), Thyanta pallidovirens (redshouldered stinkbug), Phthorimaea operculella (potato tuberworm), Helicoverpa zea (tomato fruitworm), Keiferia lycopersicella (tomato pinworm), Limonius wireworms, Manduca sexta (tobacco hornworm), and M. quinquemaculata (tomato hornworm), Liriomyza sativae, L. trifolli, L. huldobrensis (leafminer), Drosophilla melanogaster, D. yakuba, D. pseudoobscura, D. simulans), Carabus granulatus, Chironomus tentanus, Ctenocephalides felis (cat flea), Diaprepes abbreviates (root weevil), Ips pini (pine engraver), Tribolium castaneum (red floor beetle), Glossina morsitans (tsetse fly), Anopheles gambiae (malaria mosquito), Helicoverpa armigera (African Bollworm), Acyrthosiphon pisum (pea aphid), Apis melifera (honey bee), Homalodisca coagulate (glassy-winged sharpshooter), Aedes aegypti (yellow fever mosquito), Bombyx mori (silkworm)), Locusta migratoria (migratory locust), Boophilus microlus (cattle tick), Acanthoscurria gomesiane (red-haired chololate bird eater). Diploptera punctala (pacific beetle cockroach), Heliconius erato (red passion flower butterfly), H. melpomene (postman butterfly)), Curculio glandium (acorn weevil), Plutella xylostella (diamontback moth), Amblyomma variegatum (cattle tick), Anteraea yamamai (silkmoth), and Armigeres subalbatus.