Modified Nucleic Acid Aptamer Generation via Segmentation
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Solution Overview
Problem
The standard aptamer generation process, SELEX, often fails to yield aptamers with comparable affinity and specificity to antibodies, and natural nucleotides offer a limited repertoire of chemical interactions, being susceptible to nuclease degradation which limits their in vivo half-life, making it challenging to incorporate modified nucleotides for enhanced aptamer discovery.
Innovation Solution
A method for generating a pool of modified nucleic acid agents with unlimited chemical diversity by immobilizing partially double-stranded candidate nucleic acid agents to a solid support, extending the forward strand with nucleotide polymerization, and incorporating modified nucleotides, allowing for simultaneous generation and amplification of modified aptamers in a clonal manner.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Adaptability or versatility
If modified nucleotides are incorporated into aptamer discovery, then chemical diversity and affinity are improved, but amplification capability deteriorates
Solution Approach 1:
The nucleic acid agent is segmented into two functional parts: a modified nucleic acid agent providing chemical diversity and a natural nucleic acid agent providing amplification capability. This segmentation allows each component to perform its specialized function without compromising the other.
Solution Approach 2:
The natural nucleic acid agent acts as an intermediary that bridges the modified nucleic acid agent and the amplification system. It contains the same nucleotide sequence as the modified agent but lacks modifications, enabling it to serve as a functional template for PCR amplification while the modified agent provides the desired chemical properties.
2Adaptability or versatility
If chemical synthesis is used to produce modified nucleic acid agents, then chemical diversity is improved, but production cost and labor intensity increase
Solution Approach 1:
Instead of chemically synthesizing each modified nucleic acid agent individually, the method creates a natural nucleic acid copy that can be amplified through PCR. This copying approach allows multiple copies to be generated enzymatically rather than synthetically, dramatically reducing cost and labor.
Solution Approach 2:
The method changes the production parameter from chemical synthesis to enzymatic amplification. By using PCR with the natural nucleic acid agent as template, the system transitions from a chemically intensive process to a biologically driven process that is more scalable and cost-effective.
3Ease of manufacture
If natural nucleotides are used, then amplification capability is maintained, but in vivo half-life and stability deteriorate
Solution Approach 1:
The system segments the functional requirements by placing the modified nucleotides (for stability) in one agent and the amplification template (for reproduction) in another. This allows the modified agent to resist nuclease degradation in vivo while the natural agent ensures amplification capability.
Solution Approach 2:
The natural nucleic acid agent serves as a copy that can be amplified and then used to produce the modified aptamer. This copying mechanism ensures that the stable modified version can be continuously regenerated without compromising its stability properties.
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
Enables the cost-effective generation of aptamers with superior affinity and specificity, overcoming the limitations of natural nucleotides and facilitating the development of high-performance aptamers for diverse applications.
Implementation Method 1
extending the forward strand with nucleotide polymerization using the corresponding reverse strand as a template, and at least one modified nucleotide is incorporated into the forward strand during extension
Implementation Method 2
each of the partially double-stranded candidate nucleic acid agents comprises a forward strand and a reverse strand longer than the forward strand, wherein the forward and reverse strands associate with each other at least partially via base-paring
Data Source
Figure 1a~1f
Figure 2a~2g
Figure 3a~3e
AI summary
The present disclosure provides compositions, methods and systems for generating nucleic acid agents having a desired property, such as a property for specifically binding to a target. More specifically, the present disclosure provides compositions, methods and systems for generating a pool of modified members comprising modified nucleic acid agents with an unlimited range of chemical diversity.