Alternative Nucleotide Flow Ordering in Sequencing-by-Synthesis
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Solution Overview
Problem
Sequencing-by-synthesis techniques face challenges with incomplete extension (IE) and carry forward (CF) errors, leading to signal loss and reduced signal-to-noise ratio as sequencing progresses.
Innovation Solution
The use of an alternative flow ordering for nucleotides in sequencing-by-synthesis reactions, where the ordering is not a continuous repeat of the same sequence, helps reduce and correct the loss of phasic synchrony caused by IE and CF errors.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional continuous repeat ordering of dNTPs is used, then the sequencing process is simple and efficient, but incomplete extension and carry forward errors accumulate leading to signal loss and reduced signal-to-noise ratio
Solution Approach 1:
The patent applies dynamics by transitioning from a static, continuous repeat dNTP ordering to a dynamic, variable ordering pattern. The flow ordering is changed based on detected errors (IE and CF) to adaptively correct dephasing in real-time, allowing the system to respond to and compensate for sequencing errors as they occur.
Solution Approach 2:
The patent implements feedback mechanisms where the system monitors sequencing signals for incomplete extension and carry forward errors, then uses this information to adjust the dNTP flow ordering. This closed-loop approach allows the system to detect errors and apply corrective ordering patterns to maintain phasic synchrony and signal quality.
2Measurement precision
If alternative flow ordering is implemented to correct IE and CF errors, then phasic synchrony is improved and signal quality is maintained, but the complexity of the sequencing system increases
Solution Approach 1:
The patent applies parameter changes by modifying the temporal parameters of dNTP delivery - specifically the ordering sequence and timing of nucleotide flows. By varying these temporal parameters based on error detection, the system corrects phasic synchrony without changing the fundamental chemistry or hardware architecture, thereby improving signal detection precision with minimal increase in system complexity.
Data Source
AI summary
A method of sequencing a polynucleotide strand can include providing the polynucleotide strand with a primer annealed thereto and a polymerase operably bound to the polynucleotide strand; successively exposing the polynucleotide strand to the flow of four different dNTPs according to a first predetermined ordering; and successively exposing the polynucleotide strand to the flow of four different dNTPs according to a second predetermined ordering, wherein the second predetermined ordering is different from the first predetermined ordering.