Models for analyzing data from sequencing-by-synthesis operations
First Claim
1. A method of estimating a number of nucleotide incorporations when sequencing a polynucleotide strand, comprising:
- flowing a series of nucleotide flows onto a reactor array comprising a plurality of chemFET sensors and having multiple reaction confinement regions, one or more copies of the polynucleotide strand being located in a loaded reaction confinement region of the reactor array, wherein the one or more copies of the polynucleotide strand are attached to a substrate particle;
receiving output signals from the plurality of chemFET sensors of the reactor array, the output signals being generated in response to chemical reactions between the one or more copies of the polynucleotide strand and the series of nucleotide flows;
determining a background signal for the loaded reaction confinement region using a model of an output signal from the loaded reaction confinement region, the model of the output signal from the loaded reaction confinement region being a linear combination of a function of a first signal component and a function of a second signal component, wherein determining the background signal comprises fitting the model to the received output signals based on at least a ratio parameter of the function of the second signal component, wherein;
the second signal component is generated based on an output signal from an unloaded reaction confinement region of the reactor array, andthe function of the second signal component comprises (i) a rate parameter that varies mathematically with a rate of change in an amount of hydrogen ions in the loaded reaction confinement region, and (ii) the ratio parameter, which ratio parameter varies mathematically with a ratio of a rate of change in the amount of hydrogen ions in the unloaded reaction confinement region relative to the rate of change in the amount of hydrogen ions in the loaded reaction confinement region;
determining an incorporation signal from the loaded reaction confinement region using the model of the output signal from the loaded reaction confinement region, wherein determining the incorporation signal further comprises fitting the model to at least some of the received output signals based on one or more parameters of the function of the first signal component; and
determining an estimate of a number of nucleotides incorporated into the polynucleotide strand using a method selection from (i) using a peak of the determined incorporation signal to estimate the number of nucleotides incorporated, and (ii) comparing the determined incorporation signal to a set of reference incorporation signal curves to estimate the number of nucleotides incorporated.
1 Assignment
0 Petitions
Accused Products
Abstract
Mathematical models for the analysis of signal data generated by sequencing of a polynucleotide strand using a pH-based method of detecting nucleotide incorporation(s). In an embodiment, the measured output signal from the reaction confinement region of a reactor array is mathematically modeled. The output signal may be modeled as a linear combination of one or more signal components, including a background signal component. This model is solved to determine the nucleotide incorporation signal. In another embodiment, the incorporation signal from the reaction confinement region of a reactor array is mathematically modeled.
-
Citations
29 Claims
-
1. A method of estimating a number of nucleotide incorporations when sequencing a polynucleotide strand, comprising:
-
flowing a series of nucleotide flows onto a reactor array comprising a plurality of chemFET sensors and having multiple reaction confinement regions, one or more copies of the polynucleotide strand being located in a loaded reaction confinement region of the reactor array, wherein the one or more copies of the polynucleotide strand are attached to a substrate particle; receiving output signals from the plurality of chemFET sensors of the reactor array, the output signals being generated in response to chemical reactions between the one or more copies of the polynucleotide strand and the series of nucleotide flows; determining a background signal for the loaded reaction confinement region using a model of an output signal from the loaded reaction confinement region, the model of the output signal from the loaded reaction confinement region being a linear combination of a function of a first signal component and a function of a second signal component, wherein determining the background signal comprises fitting the model to the received output signals based on at least a ratio parameter of the function of the second signal component, wherein; the second signal component is generated based on an output signal from an unloaded reaction confinement region of the reactor array, and the function of the second signal component comprises (i) a rate parameter that varies mathematically with a rate of change in an amount of hydrogen ions in the loaded reaction confinement region, and (ii) the ratio parameter, which ratio parameter varies mathematically with a ratio of a rate of change in the amount of hydrogen ions in the unloaded reaction confinement region relative to the rate of change in the amount of hydrogen ions in the loaded reaction confinement region; determining an incorporation signal from the loaded reaction confinement region using the model of the output signal from the loaded reaction confinement region, wherein determining the incorporation signal further comprises fitting the model to at least some of the received output signals based on one or more parameters of the function of the first signal component; and determining an estimate of a number of nucleotides incorporated into the polynucleotide strand using a method selection from (i) using a peak of the determined incorporation signal to estimate the number of nucleotides incorporated, and (ii) comparing the determined incorporation signal to a set of reference incorporation signal curves to estimate the number of nucleotides incorporated. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20)
-
-
21. A non-transitory machine-readable storage medium comprising instructions which, when executed by a processor, cause the processor to perform a method of estimating a number of nucleotide incorporations when sequencing a polynucleotide strand comprising:
-
receiving output signals generated in response to chemical reactions resulting from a flow of a series of nucleotides onto a reactor array comprising a plurality of chemFET sensors and having multiple reaction confinement regions, one or more copies of the polynucleotide strand being located in a loaded reaction confinement region of the reactor array, wherein the one or more copies of the polynucleotide strand are attached to a substrate particle; determining a background signal for the loaded reaction confinement region using a model of an output signal from the loaded reaction confinement region, the model of the output signal from the loaded reaction confinement region being a linear combination of a function of a first signal component and a function of a second signal component, wherein determining the background signal comprises fitting the model to the received output signals based on at least a ratio parameter of the function of the second signal component, wherein; the second signal component is generated based on an output signal from an unloaded reaction confinement region of the reactor array, and the function of the second signal comprises (i) a rate parameter that varies mathematically with a rate of change in the amount of hydrogen ions in the loaded reaction confinement region, and (ii) the ratio parameter, which ratio parameter varies mathematically with a ratio of a rate of change in the amount of hydrogen ions in the unloaded reaction confinement region relative to the rate of change in the amount of hydrogen ions in the loaded reaction confinement region; determining an incorporation signal from the loaded reaction confinement region using the model of the output signal from the loaded reaction confinement region, wherein determining the incorporation signal further comprises fitting the model to at least some of the received output signals based on one or more parameters of the function of the first signal component; determining an estimate of the number of nucleotides incorporated into the polynucleotide strand using a method selected from (i) using a peak of the determined incorporation signal to estimate the number of nucleotides incorporated, and (ii) comparing the determined incorporation signal to a set of reference incorporation signal curves to estimate the number of nucleotides incorporated; and storing the estimate of the number of nucleotides incorporated in a data storage memory.
-
-
22. A method of estimating a number of nucleotide incorporations when sequencing a polynucleotide strand, comprising:
-
flowing a nucleotide reagent onto a reactor array comprising a plurality of chemFET sensors and having multiple reaction confinement regions, one or more copies of the polynucleotide strand being located in a loaded reaction confinement region of the reactor array; receiving signal data from the plurality of chemFET sensors of the reactor array, the signal data being generated in response to chemical reactions between the one or more copies of the polynucleotide strand and nucleotides from the flowing of the nucleotide reagent; processing the signal data from the loaded reaction confinement region by subtracting a background signal to obtain incorporation signal data; fitting an incorporation signal model comprising a system of non-linear differential equations to the incorporation signal data obtained from the processing, the system of non-linear differential equations comprising an equation mathematically representing a rate of change in a nucleotide concentration in the loaded reaction confinement region and an equation mathematically representing an amount of active polymerase in the loaded reaction confinement region; and determining, from the fit of the incorporation signal model, an estimate of a number of nucleotide incorporations resulting from the flowing of the nucleotide reagent. - View Dependent Claims (23, 24, 25, 26, 27, 28, 29)
-
Specification