System and Method for Correcting Primer Extension Errors in Nucleic Acid Sequence Data

• US 20100192032A1
• Filed: 02/15/2007
• Published: 07/29/2010
• Est. Priority Date: 02/16/2006
• Status: Active Grant

• Current Assignee: 454 Life Sciences Corporation (Roche Holding AG)
• Original Assignee: 454 Life Sciences Corporation (Roche Holding AG)
• Inventors: McDade, Keith, Simpson, John, Chen, Yi-Ju
• Granted Patent: US 8,301,394 B2
• Time in Patent Office: Days
• US Class Current: 714/746
• CPC Class Codes: C12Q 1/6848 characterised by the means for preventing contamination or increasing the specificity or sensitivity of an amplification reaction
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 2537/165 Mathematical modelling, e.g. logarithm, ratio
• CPC Class Codes: G16B 30/00 ICT specially adapted for sequence analysis involving nucleotides or amino acids
• CPC Class Codes: G16B 40/00 ICT specially adapted for biostatistics; ICT specially adapted for bioinformatics-related machine learning or data mining, e.g. knowledge discovery or pattern finding

Abstract

An embodiment of method for correcting an error associated with phasic synchrony of sequence data generated from a population of substantially identical copy of a template molecule is described that comprises (a) detecting a signal generated in response to an incorporation of one or more nucleotides in a sequencing reaction; (b) generating a value for the signal; and (c) correcting the value for the phasic synchrony error using a first parameter and a second carry forward parameter.

Claims

1. A method for correcting an error associated with phasic synchrony of sequence data generated from a population of substantially identical copies of a template molecule, comprising:

(a) detecting a signal generated in response to an incorporation of one or more nucleotides in a sequencing reaction;

(b) generating a value for the signal; and

(c) correcting the value for the phasic synchrony error using a first parameter and a second parameter.

2. The method of claim 1, further comprising:

(d) repeating steps (a)-(c) for each sequence position of a template molecule.

3. The method of claim 2, further comprising:

(e) incorporating each corrected value into a representation of the template molecule.

4. The method of claim 3, wherein:

the representation includes a flowgram.

5. The method of claim 2, wherein:

the phasic synchrony error includes an incomplete extension component and a carry forward component that are both treated as substantially constant for each sequence position of the template molecule, wherein the first parameter is representative of the incomplete extension component and the second parameter is representative of the carry forward component.

6. The method of claim 2, wherein:

the phasic synchrony error includes a carry forward component that is treated as substantially constant for each sequence position of the template molecule, wherein the second parameter is representative of the carry forward component.

7. The method of claim 1, wherein:

the signal includes light emitted in response to the incorporation of the one or more nucleotides.

8. The method of claim 7, wherein:

the light includes chemiluminescent light from the sequencing reaction.

9. The method of claim 8, wherein:

the sequencing reaction includes a pyrophosphate sequencing reaction.

10. The method of claim 7, wherein:

the light includes fluorescent light from the sequencing reaction.

11. The method of claim 10, wherein:

the sequencing reaction includes a sequencing reaction that employs reversible terminators.

12. The method of claim 1, wherein:

the value for the signal is indicative of a number of the incorporated nucleotides.

13. The method of claim 1, wherein:

a value for the first parameter and a value for the second parameter are estimated by searching for a best fit to a matrix equation for each of the first and second parameters.

14. The method of claim 13, wherein:

the estimation of the best fit for the first and second parameters include searching using an interval between test values and applying one or more approximations to a matrix construction operation at each test value, wherein the approximations provide for improved computational efficiency of the estimation.

15. A method for correcting an error associated with phasic synchrony of sequence data generated from a population of substantially identical copies of a template molecule, comprising:

(a) detecting a signal generated in response to an incorporation of one or more nucleotides in a sequencing reaction;

(b) generating a value for the signal;

(c) incorporating the value into a representation associated with a sequence of a template molecule;

(d) repeating steps (a)-(c) for each sequence position of the template molecule;

(e) correcting each value for the phasic synchrony error in the representation using a first parameter and a second parameter; and

(f) generating a corrected representation using the corrected values.

16. The method of claim 15, further comprising:

(g) iteratively repeating steps (e)-(f) using the corrected values from a previous iteration for step (e), wherein some or all of the corrected values improve in quality with each iteration.

17. The method of claim 15, wherein:

the phasic synchrony error includes an incomplete extension component and a carry forward component that are both treated as substantially constant for each sequence position of the template molecule, wherein the first parameter is representative of the incomplete extension component and the second parameter is representative of the carry forward component.

18. The method of claim 15, wherein:

the phasic synchrony error includes a carry forward component that is treated as substantially constant for each sequence position of the template molecule, wherein the second parameter is representative of the carry forward component.

19. The method of claim 15, wherein:

the signal includes light emitted in response to the incorporation of the one or more nucleotides.

20. The method of claim 19, wherein:

the light includes chemiluminescent light from the sequencing reaction.

21. The method of claim 20, wherein:

the sequencing reaction includes a pyrophosphate sequencing reaction.

22. The method of claim 19, wherein:

the light includes fluorescent light from the sequencing reaction.

23. The method of claim 22, wherein:

the sequencing reaction includes a sequencing reaction that employs reversible terminators.

24. The method of claim 15, wherein:

the value for the signal is indicative of a number of the incorporated nucleotides.

25. The method of claim 15, wherein:

a value for the first parameter and a value for the second parameter are estimated by searching for a best fit to a matrix equation for each of the first and second parameters.

26. The method of claim 25, wherein:

the estimation of the best fit for the first and second parameters include searching using an interval between test values and applying one or more approximations on a matrix construction operation at each test value, wherein the approximations provide for improved computational efficiency of the estimation.

27. The method of claim 15, wherein:

the representation and corrected representation include a flowgram.

28. A method for correcting an error associated with phasic synchrony of sequence data generated from a population of substantially identical copies of a template molecule, comprising:

(a) detecting a signal generated in response to an incorporation of one or more nucleotides in a sequencing reaction;

(b) generating a value for the signal;

(c) incorporating the value into a representation associated with a sequence of a template molecule;

(d) repeating steps (a)-(c) for each sequence position of the template molecule;

(e) dividing the representation into a plurality of sub-sets, wherein each sub-set comprises one or more sequence positions of the template molecule;

(f) estimating the synchrony error for a first parameter and a second parameter in each sub-set;

(g) correcting each value in each sub-set for the phasic synchrony error using the synchrony error estimations for the first parameter and for the second parameter for each respective sub-set; and

(h) combining the corrected sub-sets into a corrected representation using the corrected values.

29. The method of claim 28, wherein:

the phasic synchrony error includes an incomplete extension component and a carry forward component that fluctuate over a plurality of sequence positions of the template molecule, wherein the first parameter is representative of the incomplete extension component and the second parameter is representative of the carry forward component.

30. The method of claim 28, wherein:

the phasic synchrony error includes a carry forward component that fluctuates over a plurality of sequence positions of the template molecule, wherein the second parameter is representative of the carry forward component.

31. A system for correcting an error associated with phasic synchrony of sequence data generated from a population of substantially identical copies of a template molecule, comprising:

a computer comprising program code stored for execution thereon, the program code performing a method comprising;

(a) generating a value for a signal detected in response to an incorporation of one or more nucleotides in a sequencing reaction; and

(b) correcting the value for the phasic synchrony error using a first parameter and a second parameter.

32. The system of claim 31, wherein the method performed by the program code further comprises:

(c) repeating steps (a)-(b) for each sequence position of a template molecule.

33. The system of claim 32, wherein the method performed by the program code further comprises:

(d) incorporating each corrected value into a representation of the template molecule.

34. The system of claim 33, wherein the method performed by the program code further comprises:

(e) providing the representation to a user.

35. A system for correcting an error associated with phasic synchrony of sequence data generated from a population of substantially identical copies of a template molecule, comprising:

a computer comprising program code stored for execution thereon, the program code performing a method comprising;

(a) generating a value for a signal detected in response to an incorporation of one or more nucleotides in a sequencing reaction;

(b) incorporating the value into a representation associated with a sequence of a template molecule;

(c) repeating steps (a)-(b) for each sequence position of the template molecule;

(d) correcting each value for the phasic synchrony error in the representation using a first parameter and a second parameter; and

(e) generating a corrected representation using the corrected values

36. The system of claim 35, wherein the method performed by the program code further comprises:

(f) iteratively repeating steps (d)-(e) using the corrected values from a previous iteration for step (d), wherein some or all of the corrected values improve in quality with each iteration.

37. The system of claim 36, wherein:

the step of iteratively repeating is responsive to a user selection of a number of iterations to perform.

38. The system of claim 35, wherein the method performed by the program code further comprises:

(f) providing the corrected representation to a user.

39. A system for correcting an error associated with phasic synchrony of sequence data generated from a population of substantially identical copies of a template molecule, comprising:

a computer comprising program code stored for execution thereon, the program code performing a method comprising;

(a) generating a value for a signal detected in response to an incorporation of one or more nucleotides in a sequencing reaction;

(b) incorporating the value into a representation associated with a sequence of a template molecule;

(c) repeating steps (a)-(c) for each sequence position of the template molecule;

(d) dividing the representation into a plurality of sub-sets, wherein each sub-set comprises one or more sequence positions of the template molecule;

(e) estimating the synchrony error for a first parameter and a second parameter in each sub-set;

(f) correcting each value in each sub-set for the phasic synchrony error using the synchrony error estimations for the first parameter and for the second parameter for each respective sub-set; and

(g) combining the corrected sub-sets into a corrected representation using the corrected values.