Segmented Nucleic Acid Sequencing for Paired-End Turnaround
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Solution Overview
Problem
Existing nucleic acid sequencing technologies lack flexibility and efficiency in sequencing strategies, requiring cumbersome steps for synthetic strand removal and sequence turnaround in paired end sequencing.
Innovation Solution
A method of nucleic acid sequencing that synthesizes the sequencing synthetic strand in segments and links it to form a whole synthetic strand, using a solid support with nucleic acid linkers and single-stranded template nucleotides, allowing immobilization by the 5′ or 3′ end, and employing reversible terminators for partial sequencing and fluorescent probes for index detection.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Ease of operation
If traditional sequencing methods are used, then sequencing can be performed, but the process requires cumbersome steps for synthetic strand removal and sequence turnaround
Solution Approach 1:
The patent divides the sequencing process into distinct phases: first sequencing one end of the DNA fragment, then performing in-place turnaround to sequence the other end. The synthetic strand is retained and reused as a template for the return sequencing, eliminating the need for removal and re-synthesis steps. This segmentation of the sequencing process into manageable phases with in-place turnaround resolves the contradiction by simplifying operations while maintaining efficiency.
Solution Approach 2:
The patent performs preliminary bridge amplification to generate sufficient synthetic strands before sequencing begins. The first sequencing strand is synthesized and used as a template for the turnaround process, preparing the system in advance for the return sequencing without requiring additional synthetic strand removal or re-synthesis steps. This preliminary preparation resolves the contradiction by streamlining the overall process.
2Adaptability or versatility
If existing sequencing strategies are used, then sequencing can be performed, but flexibility in sequencing strategies is limited
Solution Approach 1:
The patent creates a universal sequencing platform that can perform multiple sequencing strategies including paired-end sequencing, single-end sequencing, and different read length configurations. The same bridge amplification and in-place turnaround mechanism supports various sequencing applications, making the system multi-functional and adaptable to different research needs without proportionally increasing complexity.
Solution Approach 2:
The patent implements dynamic control over the sequencing process, allowing the system to adapt between different sequencing modes (paired-end, single-end, different read lengths) based on the specific application requirements. The in-place turnaround mechanism can be activated or modified depending on whether paired-end sequencing is required, providing flexibility without requiring completely different systems for each application.
3Reliability
If synthetic strands are removed and re-synthesized in paired end sequencing, then complete sequencing is achieved, but the process becomes cumbersome and time-consuming
Solution Approach 1:
The patent employs a self-service mechanism where the first sequencing strand automatically serves as the template for the return sequencing after in-place turnaround. The synthetic strand that was synthesized for the first read becomes the template for the second read without requiring removal, degradation, or re-synthesis. This self-service approach ensures complete sequencing while eliminating cumbersome manual intervention steps.
Solution Approach 2:
The patent merges the two sequencing directions (forward and return reads) into a single continuous process on the same solid support. The in-place turnaround combines the completion of the first read with the initiation of the second read, eliminating the need for separate synthetic strand removal and re-synthesis steps. This merging of operations ensures complete sequencing while dramatically simplifying the overall process.
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 provides a more flexible sequencing method that reduces the need for synthetic strand removal and sequence turnaround, simplifies the process, and can be combined with non-sequencing index detection methods, shortening sequencing time and reducing costs.
Implementation Method 1
providing a solid support with at least two types of nucleic acid linkers and a single-stranded template nucleotide, where both ends of the single-stranded template nucleotide have sequences that are complementary to the nucleic acid linkers
Implementation Method 2
forming multiple nucleotide strands identical or complementary to the single-stranded template nucleotide on the solid support by a bridge amplification reaction
Implementation Method 3
hybridizing sequencing primers of the first region to be detected and the second region to be detected on the first sequencing strand
Implementation Method 4
sequencing in segments or extending and linking into a whole synthetic strand
Data Source
AI summary
Method of nucleic acid sequencing is provided, including: S1, providing a solid support with at least two types of nucleic acid linkers and a single-stranded template nucleotide, where both ends of the single-stranded template nucleotide have sequences that are complementary to the nucleic acid linkers and the single-stranded template nucleotide has a region to be detected, respectively, and the region to be detected includes a first region to be detected and a second region to be detected; S2, forming multiple nucleotide strands identical or complementary to the single-stranded template nucleotide on the solid support by a bridge amplification reaction, removing one of the nucleotide strands identical or complementary to the single-stranded template nucleotide; and S3, hybridizing sequencing primers of the first region to be detected and the second region to be detected on the first sequencing strand, and sequencing in segments or extending and linking into a whole synthetic strand.