Consolidated pretreatment and hydrolysis of plant biomass expressing cell wall degrading enzymes

• US 9,249,474 B2
• Filed: 03/07/2012
• Issued: 02/02/2016
• Est. Priority Date: 11/06/2009
• Status: Active Grant

• Current Assignee: Agrivida, Inc. (Nippon Soda Company Limited)
• Original Assignee: Agrivida, Inc. (Nippon Soda Company Limited)
• Inventors: Raab, R. Michael, Zhang, Dongcheng, Bougri, Oleg
• Primary Examiner(s): Page, Brent T
• Application Number:
  US13/414,627
• Publication Number:
  US 20120258503A1
• Time in Patent Office: 1,427 Days
• Field of Search: None
• US Class Current: 1/1
• CPC Class Codes: C12N 15/8216 Methods for controlling, regulating or enhancing expression of transgenes in plant cells
• CPC Class Codes: C12N 15/8246 Non-starch polysaccharides, e.g. cellulose, fructans, levans
• CPC Class Codes: C12N 9/2437 Cellulases (3.2.1.4; 3.2.1.74; 3.2.1.91; 3.2.1.150)
• CPC Class Codes: C12N 9/2445 Beta-glucosidase (3.2.1.21)
• CPC Class Codes: C12N 9/248 Xylanases
• CPC Class Codes: C12P 19/14 produced by the action of a carbohydrase (EC 3.2.x), e.g. by alpha-amylase , e.g. by cellulase, hemicellulase
• CPC Class Codes: C12Y 302/01004 Cellulase (3.2.1.4), i.e. endo-1,4-beta-glucanase
• CPC Class Codes: C12Y 302/01008 Endo-1,4-beta-xylanase (3.2.1.8)
• CPC Class Codes: C12Y 302/01021 Beta-glucosidase (3.2.1.21)
• CPC Class Codes: Y02E 50/10 Biofuels, e.g. bio-diesel
• CPC Class Codes: Y02E 50/30 Fuel from waste, e.g. synthetic alcohol or diesel

Abstract

Methods for consolidated pretreatment and hydrolysis of genetically engineered plants expressing cell wall degrading enzymes are provided. Expression cassettes and vectors for making transgenic plants are described. Plants engineered to express one or more cell wall degrading enzymes using expression cassettes and vectors of the invention are also provided.

Claims

1. An engineered plant including a first polynucleotide sequence encoding an amino acid sequence with at least 90% identity to a first reference sequence selected from the group consisting of:

SEQ ID NO;

7 and SEQ ID NO;

8.

2. The engineered plant of claim 1 further including a second polynucleotide sequence encoding an amino acid sequence with at least 90% identity to a second reference sequence selected from the group consisting of:

SEQ ID NO;

1, SEQ ID NO;

2, SEQ ID NO;

3, SEQ ID NO;

4, SEQ ID NO;

5, SEQ ID NO;

6, SEQ ID NO;

7, SEQ ID NO;

8, SEQ ID NO;

9, SEQ ID NO;

10, SEQ ID NO;

11, and SEQ ID;

12, wherein the SEQ ID NO selected as the second reference sequence is different than the SEQ ID NO selected as the first reference sequence.

3. The engineered plant of claim 2 further comprising a third polynucleotide sequence encoding an amino acid sequence with at least 90% identity to a third reference sequence selected from the group consisting of SEQ ID NO:

1, SEQ ID NO;

2, SEQ ID NO;

3, SEQ ID NO;

4, SEQ ID NO;

5, SEQ ID NO;

6, SEQ ID NO;

7, SEQ ID NO;

8, SEQ ID NO;

9, SEQ ID NO;

10, SEQ ID NO;

11, and SEQ ID;

12, wherein the SEQ ID NO selected as the third reference sequence is different than the SEQ ID NO selected as the first reference sequence and different than the SEQ ID NO selected as the second reference sequence.

4. The engineered plant of claim 1, wherein the first polynucleotide sequence, the second polynucleotide sequence, or the third polynucleotide sequence further includes a targeting polynucleotide sequence encoding a targeting peptide selected from the group consisting of:

an amyloplast targeting signal, a cell wall targeting peptide, a mitochondrial targeting peptide, a cytosol localization signal, a chloroplast targeting signal, a nuclear targeting peptide, and a vacuole targeting peptide.

5. The engineered plant of claim 4, wherein the targeting peptide is selected from the group consisting of:

SEQ ID NO;

13, SEQ ID NO;

14, SEQ ID NO;

15, SEQ ID NO;

16, SEQ ID NO;

17, SEQ ID NO;

22, SEQ ID NO;

23, and SEQ ID NO;

24.

6. The engineered plant of claim 1, wherein the engineered plant is selected from the group consisting of:

corn, sugar cane, sugar beet, sorghum, switchgrass, miscanthus, eucalyptus, willow and poplar.

7. The engineered plant of claim 1, wherein the first polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the first reference sequence of SEQ ID NO:

7.

8. The engineered plant of claim 2, wherein the first polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the first reference sequence of SEQ ID NO:

7 and the second polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the second reference sequence of SEQ ID NO;

8.

9. The engineered plant of claim 1, wherein the first polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the first reference sequence of SEQ ID NO:

8.

10. The engineered plant of claim 2, wherein the second polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the second reference sequence of SEQ ID NO:

1.

11. The engineered plant of claim 2, wherein the second polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the second reference sequence of SEQ ID NO:

2.

12. The engineered plant of claim 2, wherein the second polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the second reference sequence of SEQ ID NO:

3.

13. The engineered plant of claim 2, wherein the second polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the second reference sequence of SEQ ID NO:

4.

14. The engineered plant of claim 2, wherein the second polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the second reference sequence of SEQ ID NO:

5.

15. The engineered plant of claim 2, wherein the second polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the second reference sequence of SEQ ID NO:

6.

16. The engineered plant of claim 2, wherein the second polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the second reference sequence of SEQ ID NO:

9.

17. The engineered plant of claim 2, wherein the second polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the second reference sequence of SEQ ID NO:

10.

18. The engineered plant of claim 2, wherein the second polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the second reference sequence of SEQ ID NO:

11.

19. The engineered plant of claim 2, wherein the second polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the second reference sequence of SEQ ID:

12.

20. The engineered plant of claim 3, wherein the third polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the third reference sequence of SEQ ID NO:

1.

21. The engineered plant of claim 3, wherein the third polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the third reference sequence of SEQ ID NO:

2.

22. The engineered plant of claim 3, wherein the third polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the third reference sequence of SEQ ID NO:

3.

23. The engineered plant of claim 3, wherein the third polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the third reference sequence of SEQ ID NO:

4.

24. The engineered plant of claim 3, wherein the third polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the third reference sequence of SEQ ID NO:

5.

25. The engineered plant of claim 3, wherein the third polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the third reference sequence of SEQ ID NO:

6.

26. The engineered plant of claim 3, wherein the third polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the third reference sequence of SEQ ID NO:

7.

27. The engineered plant of claim 3, wherein the third polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the third reference sequence of SEQ ID NO:

8.

28. The engineered plant of claim 3, wherein the third polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the third reference sequence of SEQ ID NO:

9.

29. The engineered plant of claim 3, wherein the third polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the third reference sequence of SEQ ID NO:

10.

30. The engineered plant of claim 3, wherein the third polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the third reference sequence of SEQ ID NO:

11.

31. The engineered plant of claim 3, wherein the third polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the third reference sequence of SEQ ID:

12.

32. The engineered plant of claim 3, wherein the first polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the first reference sequence of SEQ ID NO:

7, the second polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the second reference sequence of SEQ ID NO;

4, and the third polynucleotide sequence encodes the amino acid with at least 90% identity to the third reference sequence of SEQ ID NO;

5.

33. The engineered plant of claim 32, wherein the first polynucleotide sequence encodes the amino acid of SEQ ID NO:

7, the second polynucleotide sequence encodes the amino acid sequence of SEQ ID NO;

4, and the third polynucleotide sequence encodes the amino acid of SEQ ID NO;

5.

34. The engineered plant of claim 3, wherein the first polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the first reference sequence of SEQ ID NO:

8, the second polynucleotide sequence encodes the amino acid sequence with at least 90% identity to the second reference sequence of SEQ ID NO;

4, and the third polynucleotide sequence encodes the amino acid with at least 90% identity to the third reference sequence of SEQ ID NO;

5.

35. The engineered plant of claim 34, wherein the first polynucleotide sequence encodes the amino acid of SEQ ID NO:

8, the second polynucleotide sequence encodes the amino acid sequence of SEQ ID NO;

4, and the third polynucleotide sequence encodes the amino acid of SEQ ID NO;

5.