Patent
The invention provides nucleic acids and polypeptides for a nucleic acid polymerase from a thermophilic organism, Thermus scotoductus . The invention also provides methods for using these nucleic acids and polypeptides.
1. An isolated derivative nucleic acid polymerase comprising an amino acid sequence having at least 95% sequence identity to SEQ ID NO:14, wherein said amino acid sequence comprises an Asp in place of Gly at a position corresponding to amino acid position 46 of SEQ ID NO:13, wherein said amino acid sequence further comprises an Arg at a position corresponding to amino acid position 250 of SEQ ID NO:13, and wherein said derivative nucleic acid polymerase has decreased 5′-3′ exonuclease activity compared to a wild type nucleic acid polymerase comprising SEQ ID NO:14.
2. The derivative nucleic acid polymerase according to claim 1 , wherein said derivative nucleic acid polymerase is thermostable.
3. The derivative nucleic acid polymerase according to claim 1 , wherein said amino acid sequence further comprises a Tyr in place of Phe at a position corresponding to amino acid position 668 of SEQ ID NO:13.
4. The derivative nucleic acid polymerase according to claim 1 , wherein said amino acid sequence comprises Val and Arg at positions corresponding to amino acid positions 249 and 250, respectively, of SEQ ID NO:13.
5. The derivative nucleic acid polymerase according to claim 1 , wherein said amino acid sequence comprises Arg and Thr at positions corresponding to amino acid positions 250 and 251, respectively, of SEQ ID NO:13.
6. The derivative nucleic acid polymerase according to claim 1 , wherein said amino acid sequence comprises Val, Arg, and Thr at positions corresponding to amino acid positions 249-251, respectively, of SEQ ID NO:13.
7. An isolated derivative nucleic acid polymerase comprising an amino acid sequence having at least 95% sequence identity to SEQ ID NO:14, wherein said amino acid sequence comprises an Asp in place of Gly at a position corresponding to amino acid position 46 of SEQ ID NO:13, wherein said polymerase further comprises a Pro at a position corresponding to amino acid position 254 of SEQ ID NO:13, and wherein said nucleic acid polymerase has decreased 5′-3′ exonuclease activity compared to a wild type nucleic acid polymerase comprising SEQ ID NO:14.
8. The derivative nucleic acid polymerase according to claim 7 , wherein said derivative nucleic acid polymerase is thermostable.
9. The derivative nucleic acid polymerase according to claim 7 , wherein said amino acid sequence further comprises a Tyr in place of Phe at a position corresponding to amino acid position 668 of SEQ ID NO:13.
10. The derivative nucleic acid polymerase according to claim 7 , wherein said amino acid sequence comprises Leu and Pro at positions corresponding to amino acid positions 253 and 254, respectively, of SEQ ID NO:13.
11. The derivative nucleic acid polymerase according to claim 7 , wherein said amino acid sequence comprises Pro and Leu at positions corresponding to amino acid position 254 and 255, respectively, of SEQ ID NO:13.
12. The derivative nucleic acid polymerase according to claim 7 , wherein said amino acid sequence comprises Leu, Pro, and Leu at positions corresponding to amino acid positions 253-255, respectively, of SEQ ID NO:13.
13. An isolated derivative nucleic acid polymerase comprising an amino acid sequence having at least 95% sequence identity to SEQ ID NO:14, wherein said amino acid sequence comprises an Asp in place of Gly at a position corresponding to amino acid position 46 of SEQ ID NO:13, wherein said polymerase further comprises a Leu at a position corresponding to amino acid position 471 of SEQ ID NO:13, and wherein said nucleic acid polymerase has decreased 5′-3′ exonuclease activity compared to a wild type nucleic acid polymerase comprising SEQ ID NO:14.
14. The derivative nucleic acid polymerase according to claim 13 , wherein said derivative nucleic acid polymerase is thermostable.
15. The derivative nucleic acid polymerase of claim 13 , wherein said amino acid sequence further comprises a Tyr in place of Phe at a position corresponding to position 668 of SEQ ID NO:13.
16. The derivative nucleic acid polymerase according to claim 13 , wherein said amino acid sequence comprises Arg and Leu at positions corresponding to amino acid positions 470 and 471, respectively, of SEQ ID NO:13.
17. The derivative nucleic acid polymerase according to claim 13 , wherein said amino acid sequence comprises Leu and Glu at positions corresponding to amino acid positions 471 and 472, respectively, of SEQ ID NO:13.
18. The derivative nucleic acid polymerase according to claim 13 , wherein said amino acid sequence comprises Arg, Leu, and Glu at positions corresponding to amino acid positions 470-472, respectively, of SEQ ID NO:13.