Predictive Model for Use in Sequencing-by-Synthesis

Abstract

A method of obtaining a more accurate estimate of a signal correction parameter(s) in sequencing-by-synthesis operations, such as incomplete extension rates, carry forward rates, and/or signal droop rates. The sequencing operation produces signal data. A model is constructed to simulate a population of template strands as it undergoes the sequencing process and becomes divided into different phase-states as the sequencing-by-synthesis progresses. For example, the model may be a phase-state model. The output from the model is used to adjust the signal correction parameter(s). For example, the model may be fitted to the signal data. This fitting results in a more accurate estimate of the signal correction parameter(s). In another embodiment, the signal droop rate is modeled as a decaying function and this decaying function is fitted to the signal data to obtain an improved estimate of the signal droop rate.

1 . A method of sequencing a polynucleotide strand, comprising:
 (a) receiving signal data relating to chemical reactions resulting from a flow of nucleotides;   (b) determining, using the signal data, sequence information of at least a portion of the polynucleotide strand;   (c) constructing a phase-state model for a set of flows that encompasses the sequence information, wherein the model includes a signal correction parameter and wherein the model is stored in a machine-readable memory;   (d) calculating, using the phase-state model, predicted signals resulting from the set of nucleotide flows;   (e) comparing the predicted signals to the signal data;   (f) adjusting the signal correction parameter of the phase-state model based on the comparison of the predicted signals to the signal data; and   (g) storing the adjusted signal correction parameter in the memory.   
     
     
         2 . The method of  claim 1 , wherein the nucleotides are flowed onto an array having multiple wells, wherein the polynucleotide strand is contained in a first well of the array, and further comprising:
 obtaining signal data relating to chemical reactions in a plurality of other wells within a region around the first well; and   performing steps (b) through (g) for each of the obtained signal data from the plurality of other wells to obtain multiple adjusted signal correction parameters.   
     
     
         3 . The method of  claim 2 , further comprising calculating a region-wide estimate of the signal correction parameter using the multiple adjusted signal correction parameters. 
     
     
         4 . The method of  claim 3 , wherein the comparing step comprises calculating a fitting metric that measures the fit between the predicted signals and the signal data from at least some of the plurality of wells. 
     
     
         5 . The method of  claim 4 , wherein the fitting metric measures the fit between the predicted signals and the signal data from less than all of the plurality of wells; and
 wherein the region-wide estimate excludes adjusted signal correction parameters from wells that produce a fitting metric exceeding a predetermined threshold.   
     
     
         6 . The method of  claim 3 , further comprising performing a base calling analysis of the signal data from multiple wells within the region using the region-wide estimate of the signal correction parameter. 
     
     
         7 . The method of  claim 1 , further comprising repeating steps (c) through (g) using the adjusted signal correction parameter. 
     
     
         8 . The method of  claim 7 , wherein use of the adjusted signal correction parameter improves the fit between the signal data and the predicted signals. 
     
     
         9 . The method of  claim 1 , wherein the phase-state model includes two or more signal correction parameters, including a carry forward rate and an incomplete extension rate. 
     
     
         10 . The method of  claim 1 , wherein the comparing step comprises calculating a fitting metric that measures a fit between the predicted signals and the signal data. 
     
     
         11 . The method of  claim 10 , wherein the adjusting step comprises determining a value of the signal correction parameter that optimizes the fitting metric. 
     
     
         12 . The method of  claim 11 , wherein the fitting metric is calculated using only nucleotide flows that result in nucleotide non-incorporation or single nucleotide incorporations. 
     
     
         13 . The method of  claim 1 , further comprising performing a base calling analysis of the signal data using the adjusted signal correction parameter. 
     
     
         14 . The method of  claim 1 , wherein the set of nucleotide flows is a first set of nucleotide flows and the sequence information is a first sequence information, and further comprising:
 applying the phase-state model using the adjusted signal correction parameter;   calculating, using the phase-state model, predicted signals resulting from a second set of nucleotide flows that includes nucleotide flows that are not in the first set of nucleotide flows;   making base calls by comparing the signal data to the predicted signals; and   obtaining a second sequence information about the polynucleotide strand, wherein the second sequence information includes sequence information not contained in the first sequence information.   
     
     
         15 . The method of  claim 14 , further comprising repeating steps (d) through (g) using the second sequence information to obtain a further adjusted signal correction parameter. 
     
     
         16 . The method of  claim 3 , wherein the region is a first region and wherein the phase-state model is adjusted for a signal droop rate that is obtained by a method comprising:
 receiving signal data relating to chemical reactions in a plurality of wells within a second region of the array, wherein the plurality of wells includes the well containing the polynucleotide strand, wherein the second region is the same or different from the first region;   calculating a set of averaged signal values from the signal data; and   determining a region-wide signal droop rate by fitting a signal decay function to the set of averaged signal values.   
     
     
         17 - 19 . (canceled) 
     
     
         20 . A method of sequencing a polynucleotide strand contained in a well of an array having multiple wells, comprising:
 receiving signal data relating to chemical reactions in a plurality of wells within a region of the array resulting from a flow of nucleotides to the array, wherein the plurality of wells includes the well containing the polynucleotide strand;   calculating a set of averaged signal values from the signal data, wherein the set of averaged signal values are stored in a machine-readable memory;   determining a region-wide signal droop rate by fitting a signal decay function to the set of averaged signal values; and   storing the region-wide signal droop rate in the memory.