Compositions and methods for the suppression of target polynucleotides from lepidoptera

Patent metadata

Field	Value
Publication number	US-9617559-B2
Application number	US-201414501240-A
Country	US
Kind code	B2
Filing date	Sep 30, 2014
Priority date	Jan 17, 2008
Publication date	Apr 11, 2017
Grant date	Apr 11, 2017

Abstract

Methods and compositions are provided which employ a silencing element that, when ingested by a pest, such as a pest from the Lepidoptera order, they are capable of decreasing the expression of a target sequence in the pest. Further provided are silencing elements which when ingested by the pest decrease the level of the target polypeptide and thereby control the pest. In specific embodiment, the pest is Spodoptera frugiperda . Plants, plant part, bacteria and other host cells comprising the silencing elements or an active variant or fragment thereof of the invention are also provided.

First claim

The invention claimed is:

1. A plant cell having stably incorporated into its genome a heterologous polynucleotide encoding a double stranded RNA, wherein said double stranded RNA comprises a sense and an antisense strand, wherein the antisense strand comprises at least 20 consecutive nucleotides fully complementary to the target gene sequence set forth in SEQ ID NO: 6, and wherein the sense strand is fully complementary to the antisense strand,
wherein said double stranded RNA, when ingested by a pest from the Lepidoptera order, controls the pest from the Lepidoptera order.

2. The plant cell of claim 1 , wherein said pest comprises Spodoptera frugiperda.

3. The plant cell of claim 1 , wherein said double stranded RNA comprises
a) the sense and antisense sequence of the sequence set forth in SEQ ID NO: 93, 96, or 99; or
b) the sequences set forth in SEQ ID NO: 94, and 95, 97, and 98, or 100 and 101.

4. The plant cell of claim 1 , wherein said double stranded RNA comprises a hairpin RNA.

5. The plant cell of claim 4 , wherein said double stranded RNA comprises a first segment, a second segment, and a third segment, wherein
a) said first segment comprises at least 20 consecutive nucleotides fully complementary to the target sequence set forth in SEQ ID NO: 6;
b) said second segment comprises a loop of sufficient length to allow the double stranded RNA to be transcribed as a hairpin RNA; and,
c) said third segment comprises at least about 20 consecutive nucleotides fully complementary to the first segment.

6. The plant cell of claim 1 , wherein said polynucleotide is operably linked to a heterologous promoter.

7. The plant cell of claim 1 , wherein said plant cell has stably incorporated into its genome a second polynucleotide comprising a suppressor enhancer element comprising the target pest sequence or an active variant or fragment thereof, wherein the combined expression of the double stranded RNA and the suppressor enhancer element increases the concentration of an inhibitory RNAi specific for the pest target sequence in said plant cell.

8. The plant cell of claim 1 , wherein said plant cell is from a monocot.

9. The plant cell of claim 8 , wherein said monocot is maize, barley, millet, wheat or rice.

10. The plant cell of claim 1 , wherein said plant cell is from a dicot.

11. The plant cell of claim 10 , wherein said dicot is soybean, canola, alfalfa, sunflower, safflower, tobacco, Arabidopsis , or cotton.

12. A plant or plant part comprising a plant cell of claim 1 .

13. The plant or plant part of claim 7 , wherein the combined expression of said double stranded RNA and the suppressor enhancer element increases the concentration of an inhibitory RNA specific for the pest target sequence in the phloem of said plant or plant part.

14. A transgenic seed from the plant of claim 12 .

15. A method for controlling Lepidoptera comprising feeding to a Lepidoptera a composition comprising a double stranded RNA, wherein the double stranded RNA comprises a sense and an antisense strand, wherein the antisense strand comprises at least 20 consecutive nucleotides fully complementary to the target gene sequence set forth in SEQ ID NO: 6, and wherein the sense strand is fully complementary to the antisense strand,
wherein said double stranded RNA, when ingested by said Lepidoptera controls the Lepidoptera.

16. The method of claim 15 , wherein said composition comprises a plant or plant part having stably incorporated into its genome a polynucleotide encoding said double stranded RNA.

17. The method of claim 15 , wherein said pest comprises Spodoptera frugiperda.

18. The method of claim 15 , wherein said double stranded RNA comprises:
a) the sense and antisense sequence of the sequence set forth in SEQ ID NO: 93, 96, or 99; or
b) the sequences set forth in SEQ ID NO: 94 and 95, 97 and 98, or 100 and 101.

19. The method of claim 15 , wherein said double stranded RNA comprises a hairpin RNA.

20. The method of claim 19 wherein said double stranded RNA comprises a first segment, a second segment, and a third segment, wherein
a) said first segment comprises at least 20 consecutive nucleotides fully complementary to the target polynucleotide;
b) said second segment comprises a loop of sufficient length to allow the double stranded RNA to be transcribed as a hairpin RNA; and,
c) said third segment comprises at least 20 consecutive nucleotides fully complementary to the first segment.

21. The method of claim 16 , wherein said plant or plant part has stably incorporated into its genome a second polynucleotide comprising a suppressor enhancer element comprising the target pest sequence or an active variant or fragment thereof, wherein the combined expression of the double stranded RNA and the suppressor enhancer element increases the concentration of an inhibitory RNAi specific for the pest target sequence in said plant cell.

22. The method of claim 15 , wherein said plant is a monocot.

23. The method of claim 22 , wherein said monocot is maize, barley, millet, wheat or rice.

24. The method of claim 15 , wherein said plant is a dicot.

25. The method of claim 24 , wherein said plant is soybean, canola, alfalfa, sunflower, safflower, tobacco, Arabidopsis , or cotton.

Assignees

• Pioneer Hi Bred Int
• Du Pont
• Pioneer Hi Bred Int

Inventors

• Herrmann Rafael
• Lassner Michael
• Lu Albert L
• Nelson Mark Edward
• Presnail James Kevin
• Rice Janet A

Classifications

• C12N2310/14

  interfering nucleic acids [NA] · CPC title

• C12N2320/32

  Special delivery means, e.g. tissue-specific · CPC title

• C12N2310/531

  Stem-loop; Hairpin · CPC title

• C12N15/8286 Primary

  for insect resistance · CPC title

• A01N65/20

  Fabaceae or Leguminosae [Pea or Legume family], e.g. pea, lentil, soybean, clover, acacia, honey locust, derris or millettia · CPC title