Terminator Nucleotides with Ion-Sensitive FET Sensors
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Solution Overview
Problem
Current nucleic acid sequencing methods face challenges in detecting nucleotide incorporation events with limited signal magnitude and transient signals, which can reduce sequencing efficiency and run length due to stoichiometric limitations and transient nature of reaction by-products.
Innovation Solution
The use of modified nucleotides that can generate sustained non-optical signals, such as those linked with terminator or tag groups, which are detectable by sensors like ion-sensitive FETs or chemically-sensitive FETs, allowing for increased signal detection and longer sequencing runs.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If traditional optical detection methods are used for nucleotide incorporation, then the sequencing process can be performed, but the signal magnitude is limited and the signal is transient, reducing sequencing efficiency and run length
Solution Approach 1:
The patent replaces optical detection methods with electrochemical detection using ion-sensitive FET sensors. The sensor detects changes in local ion concentration near the reaction site, converting chemical information into electrical signals that are sustained and easily detectable, thereby resolving the contradiction between detection capability and signal duration.
2Productivity
If stoichiometric amounts of nucleotide are used, then the sequencing reaction proceeds, but the signal is transient and limited in magnitude due to the nature of by-product release
Solution Approach 1:
The patent introduces ion-sensitive FET sensors as intermediaries that detect local ion concentration changes near the reaction site. These sensors act as mediators between the nucleotide incorporation event and the detection system, converting transient chemical events into sustained electrical signals that can be detected with high fidelity, thereby resolving the contradiction between productivity and information retention.
3Measurement precision
If optical labels are attached to nucleotides for detection, then nucleotide incorporation can be detected, but the system complexity increases and the signal remains transient
Solution Approach 1:
The patent replaces complex optical labeling systems with electrochemical sensors that detect intrinsic ion concentration changes during nucleotide incorporation. This substitution eliminates the need for optical labels and their associated complexity while providing sustained, high-fidelity detection signals, thereby resolving the contradiction between measurement precision and device complexity.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This approach enhances the detection of nucleotide incorporation events by generating sustained signals, improving sequencing efficiency and run length by increasing the magnitude and duration of detectable signals, thereby overcoming the limitations of stoichiometric and transient signals in existing methods.
Implementation Method 1
detecting a signal (e.g., a non-optical signal) indicating the nucleotide incorporation using a sensor that is attached or operatively linked to the at least one reaction site
Implementation Method 2
detectable by sensors like ion-sensitive FETs or chemically-sensitive FETs
Data Source
AI summary
In some embodiments, the disclosure relates generally to methods, as well as related, systems, compositions, kits and apparatuses, for nucleic acid analysis that involve the use of modified nucleotides, including terminator nucleotides and/or tagged nucleotides, in a template-dependent nucleotide incorporation reaction. In some embodiments, the nucleic acid analysis can be conducted at a single reaction site, or at a plurality of reaction sites in an array of reaction sites. Optionally, the array contains a plurality of reaction sites having about 1-100 million, or about 100-250 million, or about 200-500 million, or about 500-900 million, or more reaction sites. Optionally, each reaction site is in contact with, operatively coupled, or capacitively coupled to one or more sensors that are ion-sensitive FETs (isFETs) or chemically-sensitive FETs (chemFETs) sensors. Optionally, the reaction sites are in fluid communication with each other.