ENZYME

• US 20110008848A1
• Filed: 02/13/2009
• Published: 01/13/2011
• Est. Priority Date: 02/28/2008
• Status: Active Grant

• Current Assignee: Genesys Biotech Ltd.
• Original Assignee: Genesys Logic
• Inventors: Morant, Nicholas, Clark, Duncan
• Granted Patent: US 8,986,968 B2
• Time in Patent Office: Days
• US Class Current: 435/91.2
• CPC Class Codes: C12N 9/1252 DNA-directed DNA polymerase (2.7.7.7), i.e. DNA replicase

Abstract

There is provided a polypeptide having thermostable DNA polymerase activity and comprising or consisting of an amino acid sequence with at least 55% identity to Thermodesulfatator indicus DNA polymerase I Large fragment shown in SEQ ID NO: 1 or in SEQ ID NO:32.

Claims

1. A polypeptide having thermostable DNA polymerase activity and which exhibits strand displacement activity, comprising or consisting of an amino acid sequence with at least 55% identity to Thermodesulfatator indicus DNA polymerase I Large fragment shown in SEQ ID NO:

1.

3. A polypeptide according to claim 1, which is suitable for carrying out an isothermal amplification reaction, such as loop-mediated isothermal amplification (LAMP) and/or which is suitable for carrying out a thermocycling amplification reaction, such as a polymerase chain reaction (PCR).

5. A polypeptide according to claim 1, wherein the amino acid sequence is SEQ ID NO:

32 or an amino acid sequence having at least 55% identity to SEQ ID NO;

32.

6. An isolated polypeptide according to claim 1, having thermostable DNA polymerase activity and comprising or consisting of an amino acid sequence with at least 55% identity to T. indicus DNA polymerase I as shown in SEQ ID NO:

2.

7. A polypeptide according to claim 6 wherein the amino acid sequence is SEQ ID NO:

34 or an amino acid sequence having at least 55% identity to SEQ ID NO;

34.

8. A polypeptide according to claim 6, in which the polypeptide is an isolated thermostable DNA polymerase I obtainable from T. indicus and having a molecular weight of about 103,000 Daltons, or an enzymatically active fragment thereof.

9. A polypeptide according to claim 1, having thermostable DNA polymerase activity and comprising amino acid sequence SEQ ID NO:

32 or amino acid sequence SEQ ID NO;

34.

13. A polypeptide according to claim 1, further comprising a Cren7 enhancer domain.

14. A composition comprising the polypeptide of claim 1.

15. An isolated nucleic acid encoding the polypeptide of claim 1.

24. A vector comprising the isolated nucleic acid of claim 15.

25. A host cell transformed with the nucleic acid of claim 15.

26. A kit comprising the polypeptide of claim 1, together with packaging materials therefor.

27. A method of amplifying a sequence of a target nucleic acid using a thermocycling reaction, comprising the steps of:

(1) contacting the target nucleic acid with the polypeptide of claim 1; and

(2) incubating the target nucleic acid with the polypeptide under thermocycling reaction conditions which allow amplification of the target nucleic acid.

30. A method of amplifying a sequence of a target nucleic acid using an isothermal reaction, comprising the steps of:

(1) contacting the target nucleic acid with the polypeptide of claim 1; and

(2) incubating the target nucleic acid with the polypeptide under isothermal reaction conditions which allow amplification of the target nucleic acid.

34. An isolated nucleic acid according to claim 15, having a nucleotide sequence selected from SEQ ID NO:

3, SEQ ID NO;

33, SEQ ID NO;

4, and SEQ ID NO;

35.

35. A host cell transformed with the vector of claim 24.

36. A kit comprising the composition of claim 14, together with packaging materials therefor.

37. A kit comprising the isolated nucleic acid of claim 15, together with packaging materials therefor.

38. A kit comprising the vector of claim 24, together with packaging materials therefor.

39. A kit comprising the host cell of claim 25, together with packaging materials therefor.

40. A kit comprising the host cell of claim 35, together with packaging materials therefor.

2. (canceled)

4. (canceled)

10-12. -12. (canceled)

16-23. -23. (canceled)

28. (canceled)

29. (canceled)

31-33. -33. (canceled)