Phase-protecting reagent flow orderings for use in sequencing-by-synthesis

Abstract

A system for nucleic acid sequencing includes a machine-readable memory and a processor configured to execute machine-readable instructions. The instructions, when executed by the processor, cause the system to expose template polynucleotide strands in a plurality of defined spaces of a sensor array to a series of flows of nucleotide species, the series comprising a sequence of random flows; and obtain, for each of the series of flows of nucleotide species, a signal indicative of how many nucleotide incorporations occurred for that particular flow to determine a predicted sequence of nucleotides corresponding to the template polynucleotide strands.

What is claimed is: 
     
         1 . A system for nucleic acid sequencing, comprising:
 a machine-readable memory; and   a processor configured to execute machine-readable instructions, which, when executed by the processor, cause the system to:
 expose template polynucleotide strands in a reaction array to a phase-protecting flow ordering of nucleotide species, 
 obtain, for each flow of nucleotide species, a signal indicative of how many nucleotide incorporations occurred, and 
 determine, based on the obtaining the signal for each flow of nucleotide species, a sequence of nucleotides corresponding to the template polynucleotide strands, 
   wherein the phase-protecting flow ordering:
 (a) is not a series of consecutive repetitions of a 4-flow permutation of the four different nucleotide species, and 
 (b) comprises a first flow ordering in which a first nucleotide species, a second nucleotide species, and a third nucleotide species are flowed at least twice before a fourth nucleotide species is flowed, and the fourth nucleotide species is flowed at least one following the flows of the first, second, and third nucleotide species. 
   
     
     
         2 . The system of  claim 1 , wherein the phase-protecting flow ordering further comprises a second flow ordering in which the second nucleotide species, the third nucleotide species, and the fourth nucleotide species are flowed at least twice before the first nucleotide species is flowed. 
     
     
         3 . The system of  claim 2 , wherein the phase-protecting flow ordering further comprises a third flow ordering in which the first nucleotide species, the third nucleotide species, and the fourth nucleotide species are flowed at least twice before the second nucleotide species is flowed. 
     
     
         4 . The system of  claim 1 , wherein the phase-protecting flow ordering further comprises:
 a flow of N followed immediately by a flow of X, wherein X and N represent different nucleotide species, and   immediately after the flow of X or elsewhere in the phase-protecting flow ordering, a flow of N followed immediately by a flow of Y, wherein Y represents a nucleotide species different from both X and N.   
     
     
         5 . The system of  claim 1 , wherein the phase-protecting flow ordering further comprises:
 a flow of X followed immediately by a flow of N, wherein X and N represent different nucleotide species, and   a flow of Y followed immediately by a flow of N, wherein Y represents a nucleotide species different from both X and N.   
     
     
         6 . The system of  claim 1 , wherein the first flow ordering comprises the flow of the fourth nucleotide species immediately following the flows of the first, second, and third nucleotide species 
     
     
         7 . The system of  claim 1 , further comprising a sensing device configured to detect a nucleotide incorporation and transmit the signal indicative of how many nucleotide incorporations occurred to the processor. 
     
     
         8 . The system of  claim 7 , wherein the sensing device is configured to provide a current-based output signal, or a voltage output signal, or both. 
     
     
         9 . The system of  claim 7 , wherein the sensing device comprises a chemically sensitive field effect transistor, and the chemically sensitive field effect transistor is used to detect a reactant concentration, a product concentration, an ion concentration, or any combination thereof. 
     
     
         10 . The system of  claim 7 , wherein the sensing device comprises an ion sensitive field effect transistor, and the ion sensitive field effect transistor is used to detect a hydrogen ion concentration. 
     
     
         11 . The system of  claim 1 , wherein the reaction array comprises one or more substrates exhibiting a binding affinity with the polynucleotide template strands. 
     
     
         12 . The system of  claim 11 , wherein the one or more substrates comprise a plurality of beads. 
     
     
         13 . A system for nucleic acid sequencing, comprising:
 a machine-readable memory; and   a processor configured to execute machine-readable instructions, which, when executed by the processor, cause the system to:
 expose template polynucleotide strands in a reaction array to a phase-protecting flow ordering of nucleotide species, 
 obtain, for each flow of nucleotide species, a signal indicative of how many nucleotide incorporations occurred, and 
 determine, based on the obtaining the signal for each flow of nucleotide species, a sequence of nucleotides corresponding to the template polynucleotide strands, 
   wherein the phase-protecting flow ordering:
 (a) is not a series of consecutive repetitions of a 4-flow permutation of the four different nucleotide species, and 
 (b) comprises a flow of N followed immediately by a flow of X, where X and N represent different nucleotide species, and further comprises, after the flow of X in the phase-protecting flow ordering, a flow of N followed immediately by a flow of Y, the flow of Y followed immediately by a flow of Z, where Y represents a nucleotide species different from both X and N, and where Z represents a nucleotide species different from X, N, and Y. 
   
     
     
         14 . The system of  claim 13 , further comprising a sensing device configured to detect a nucleotide incorporation and transmit the signal indicative of how many nucleotide incorporations occurred to the processor. 
     
     
         15 . The system of  claim 14 , wherein the sensing device is configured to provide a current-based output signal, or a voltage output signal, or both. 
     
     
         16 . The system of  claim 13 , wherein the sensing device comprises a chemically sensitive field effect transistor, and the chemically sensitive field effect transistor is used to detect a reactant concentration, a product concentration, an ion concentration, or any combination thereof. 
     
     
         17 . The system of  claim 13 , wherein the sensing device comprises an ion sensitive field effect transistor, and the ion sensitive field effect transistor is used to detect a hydrogen ion concentration. 
     
     
         18 . The system of  claim 13 , wherein the reaction array comprises one or more substrates exhibiting a binding affinity with the polynucleotide template strands. 
     
     
         19 . The system of  claim 13 , wherein the one or more substrates comprise a plurality of beads.