Silicon Modified Nucleotide Analogues for Sequencing Accuracy
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Solution Overview
Problem
Current DNA sequencing methods, particularly sequencing by synthesis (SBS), face challenges in accurately and efficiently incorporating modified nucleotides recognized by DNA polymerases, leading to issues such as premature termination and background signal interference due to reactive thiols and linker interactions.
Innovation Solution
Development of novel nucleotide analogues with specific divalent linkers and cleavable moieties that minimize reactive thiol formation and linker interactions, incorporating detectable labels for accurate sequencing, and using polymerase-compatible cleavable moieties to enhance sequencing accuracy.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If modified nucleotides with divalent linkers and detectable labels are used in sequencing by synthesis, then sequencing accuracy and detectability are improved, but reactive thiol formation and linker interactions cause premature termination and background signal interference
Solution Approach 1:
The patent extracts and removes the problematic thiol group from the nucleotide structure. By replacing the thiol-containing linker with a thiol-free alternative, the source of premature termination and background signal is eliminated while retaining the essential functionality of the detectable label and linker system.
Solution Approach 2:
The patent introduces a thiol-free linker as an intermediary component that connects the nucleobase to the detectable label without forming reactive thiols. This intermediary structure maintains the necessary spacing and structural integrity while avoiding the harmful chemical reactivity that causes sequencing errors.
2Productivity
If modified nucleotides with detectable labels are incorporated into growing DNA chains, then nucleotide incorporation efficiency is improved, but background signal interference occurs due to linker interactions
Solution Approach 1:
The patent converts the potentially harmful thiol group into a beneficial thiol-free structure. By removing the reactive thiol that causes background signal interference, the detectable label can function more reliably, allowing the sequencing system to achieve both high incorporation efficiency and clean signal detection.
Solution Approach 2:
The patent changes the chemical parameters of the linker structure by removing the thiol functional group. This parameter change fundamentally alters the chemical reactivity profile of the modified nucleotide, eliminating the source of background signal while preserving the detectability and incorporation efficiency functions.
3Ease of operation
If cleavable moieties are used to enable nucleotide release after incorporation, then sequencing cycle completion is improved, but reactive thiols from cleavage cause downstream complications
Solution Approach 1:
The patent extracts the problematic thiol group from the cleavable moiety structure. By designing a cleavage mechanism that does not generate free thiols, the patent eliminates the source of downstream complications such as reduced detectability and increased background signal while maintaining efficient nucleotide release.
Solution Approach 2:
The patent introduces a thiol-free cleavable linker as an intermediary that facilitates nucleotide release without generating reactive thiols. This intermediary structure enables the cleavage function while avoiding the harmful chemical byproducts that would interfere with subsequent sequencing cycles.
Data Source
AI summary
Disclosed herein, inter alia, are silicon containing cleavable linkers and compounds for use in nucleic acid sequencing.