Methods and apparatus that increase sequencing-by-binding efficiency

Patent metadata

Field	Value
Publication number	US-10161003-B2
Application number	US-201815922787-A
Country	US
Kind code	B2
Filing date	Mar 15, 2018
Priority date	Apr 25, 2017
Publication date	Dec 25, 2018
Grant date	Dec 25, 2018

Abstract

A method of determining a nucleic acid sequence that includes steps of: (a) contacting a primed template nucleic acid with a series of mixtures for forming ternary complexes, wherein each of the mixtures includes a polymerase and nucleotide cognates for at least two different base types suspected of being present at the next template position of the template nucleic acid; (b) monitoring the next template position for ternary complexes formed by the series of mixtures, wherein a signal state indicates presence or absence of ternary complex formed at the next template position by each individual mixture, thereby determining a series of signal states that encodes a base call for the next template position; and (c) decoding the series of signal states to distinguish a correct base call for the next template position from an error in the base call.

First claim

What is claimed is:

1. A method of determining a nucleic acid sequence, comprising:
(a) contacting a primed template nucleic acid with a series of mixtures for forming ternary complexes, wherein each of the mixtures comprises a polymerase and nucleotide cognates for at least two different base types suspected of being present at the next template position of the template nucleic acid, wherein the mixtures differ by the presence or absence of at least one type of nucleotide cognate;
(b) monitoring the next template position for ternary complexes formed by the series of mixtures, wherein a signal state indicates presence or absence of ternary complex formed at the next template position by each individual mixture, thereby determining a series of signal states that encodes a base call for the next template position; and
(c) decoding the series of signal states to distinguish a correct base call for the next template position from an error in the base call.

2. The method of claim 1 , wherein the series of signal states comprises an error correcting code.

3. The method of claim 2 , wherein the series of mixtures consists of three mixtures and the series of signal states is represented by three digits, each digit representing a signal state obtained from a mixture.

4. The method of claim 3 , wherein each of the signal states is represented by a binary digit, and wherein the error correcting code comprises a repetition code.

5. The method of claim 4 , further comprising correcting the invalid base call by a majority vote between the three digits.

6. The method of claim 2 , wherein the series of mixtures consists of four mixtures and the series of signal states is represented by four digits, each digit representing a signal state obtained from a mixture.

7. The method of claim 6 , wherein each of the signal states is represented by a ternary digit, wherein the error correcting code comprises a Hamming code, and wherein the Hamming distance between valid base calls is three.

8. The method of claim 7 , further comprising correcting the invalid base call to a valid base call having a code with the closest Hamming distance to the code for the invalid base call.

9. The method of claim 2 , wherein the series of mixtures consists of five mixtures and the series of signal states is represented by five digits, each digit representing a signal state obtained from a mixture.

10. The method of claim 9 , wherein each of the signal states is represented by a binary digit, wherein the error correcting code comprises a Hamming code, and wherein each valid base call differs from other valid base calls by three digits.

11. The method of claim 1 , further comprising correcting the invalid base call to a valid base call having a code with the closest Hamming distance to the code for the invalid base call.

12. The method of claim 1 , wherein the decoding of the series of signal states identifies the base call as being an invalid base call.

13. The method of claim 12 , further comprising correcting the invalid base call to make a valid base call for the next template position.

14. The method of claim 13 , wherein the correcting of the error comprises correlating a suspect signal state in the series of signal states with an aberration in step (a) or (b), and selecting the base call having the expected series of signal states with a change in the suspect signal state.

15. The method of claim 14 , wherein the aberration in step (b) is selected from the group consisting of a signal to noise ratio below a predetermined threshold, signal below a predetermined threshold, signal above a predetermined threshold, noise above a predetermined threshold, and detector malfunction.

16. The method of claim 14 , wherein the aberration in step (a) is selected from the group consisting of a fluidic delivery malfunction, temperature control malfunction, and reagent quality below a predetermined threshold.

17. The method of claim 1 , wherein the series of signal states comprises a repetition code, Hamming code, linear code or parity code.

18. The method of claim 1 , wherein each of the signal states is represented by a binary digit.

19. The method of claim 18 , wherein the binary digit comprises (i) symbols for presence and absence of a signal; (ii) symbols for signals emitted at two different wavelengths; (iii) symbols for signals having two different intensities; or (iv) symbols for signals resulting from excitation at two different wavelengths.

20. The method of claim 1 , wherein each of the signal states is represented by a ternary digit.

21. The method of claim 20 , wherein the ternary digit comprises (i) symbols for signals emitted at three different wavelengths; (ii) symbols for signals having three different intensities; or (iii) symbols for signals resulting from excitation at three different wavelengths.

22. The method of claim 1 , wherein each of the mixtures comprises nucleotide cognates for at least two and no more than three of four different base types suspected of being in the primed template nucleic acid.

23. The method of claim 1 , wherein each of the mixtures comprises nucleotide cognates for at least two and no more than two of four different base types suspected of being in the primed template nucleic acid.

24. The method of claim 1 , wherein the mixtures differ by the number or type of label attached to at least one type of nucleotide cognate.

25. The method of claim 1 , further comprising:
(d) adding a next correct nucleotide to the primer of the primed template nucleic acid after step (b), thereby producing an extended primer.

26. The method of claim 25 , further comprising repeating steps (a) through (d) for the primed template nucleic acid that comprises the extended primer.

27. The method of claim 25 , wherein the next correct nucleotide that is added to the primer is a reversibly terminated nucleotide.

28. The method of claim 27 , further comprising repeating steps (a) through (d) for the primed template nucleic acid that comprises the extended, reversibly terminated primer.

29. The method of claim 28 , further comprising (e) removing the reversible terminator moiety from the extended, reversibly terminated primer after steps (a) through (d) are repeated.

Assignees

• Omniome Inc

Inventors

• Stromberg Sean
• Vieceli John
• Vijayan Kandaswamy
• Oliphant Arnold

Classifications

• C12Q1/6874 Primary

  involving nucleic acid arrays, e.g. sequencing by hybridisation · CPC title

• C12Q1/6869

  Methods for sequencing · CPC title

• C12Q1/6832 Primary

  Enhancement of hybridisation reaction · CPC title

• C12Q1/6853

  using modified primers or templates · CPC title

• C12Q2535/122

  Massive parallel sequencing · CPC title