Solid Phase Clonal Amplification for Targeted Sequencing
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Solution Overview
Problem
Current next-generation sequencing technologies rely on random sampling methods, which are inefficient and redundant, failing to specifically target regions of interest, leading to excessive analysis of non-informative sequences.
Innovation Solution
The method involves solid phase clonal amplification of hybridized target molecules on probe arrays, allowing for locus-specific targeting and analysis by synthesizing or ligating amplified DNA sequences, reducing redundancy and enhancing sequencing efficiency.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Adaptability or versatility
If random sampling methods are used for sequencing, then all regions of the genome are covered, but redundant sequencing of non-informative regions occurs and regions of interest are not specifically targeted
Solution Approach 1:
The patent applies local quality by making different regions of the solid support have different functions: regions with probes specific to sequences of interest perform clonal amplification and sequencing, while regions without hybridized targets remain inactive. This selective activation of specific locations eliminates redundant sequencing of non-informative regions while maintaining comprehensive coverage capability.
Solution Approach 2:
The patent employs preliminary action by pre-synthesizing probes complementary to specific sequences of interest on the solid support before the sequencing process begins. These pre-positioned probes enable selective capture and amplification of target sequences, allowing the system to specifically target regions of interest without random sampling redundancy.
2Measurement precision
If clonal amplification is performed on all hybridized molecules, then sufficient signal is obtained for detection, but analysis time increases due to processing all sequences including non-informative ones
Solution Approach 1:
The patent applies local quality by making different regions of the solid support have different functions: regions with probes specific to sequences of interest perform clonal amplification and sequencing, while regions without hybridized targets remain inactive. This selective activation of specific locations eliminates redundant sequencing of non-informative regions while maintaining comprehensive coverage capability.
3Productivity
If locus-specific targeting is implemented, then sequencing efficiency is improved by reducing redundancy, but the complexity of probe design and array synthesis increases
Solution Approach 1:
The patent applies segmentation by dividing the solid support into multiple discrete regions, each containing probes for specific sequences of interest. This segmentation allows independent optimization of each probe region and enables parallel processing of multiple targets simultaneously, improving overall sequencing efficiency while managing complexity through modular design.
Solution Approach 2:
The patent employs universality by designing probes with universal adapter sequences that enable multiple functions: specific target recognition, clonal amplification priming, and sequencing initiation. This multi-functionality reduces the need for separate components and simplifies the overall system despite the complexity of locus-specific targeting.
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 enables precise targeting and amplification of specific DNA regions, reducing redundant sequencing and improving analysis efficiency by focusing on regions of interest, thereby enhancing the accuracy and productivity of nucleic acid analysis.
Implementation Method 1
hybridization of target to complementary probes on the array
Implementation Method 2
extending the probes that have hybridized to the targets, thereby obtaining extended probes that comprise copies of the targets
Data Source
AI summary
Methods are provided for multiplexed amplification of selected targets and analysis of the amplified targets. In preferred aspects the amplification and analysis take place on the same solid support and preferably in a localized area such as a bead or a feature of an array. Targets are circularized by hybridization to probes followed by ligation of the ends of the target to form a closed circle. The targets are then used as template for extension of an array bound probe resulting in extended probes having multiple copies of the target. The extended probes can then be analyzed. The methods may be used for genotyping, sequencing and analysis of copy number.