Aptamer Identification via Spatially Separated Amplification
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Solution Overview
Problem
The existing methods for identifying aptamers, such as SELEX, are time-consuming and labor-intensive, with a high risk of losing good binders due to unfavorable selection conditions and random selection processes, leading to inefficient aptamer identification and characterization.
Innovation Solution
A method involving contacting a mixture of oligonucleotides with an aptamer target structure, separating bound oligonucleotides, amplifying them spatially, labeling, sequencing, and analyzing their binding properties in parallel, allowing for the identification of aptamers with high affinity and specificity in a single selection step without the need for multiple rounds.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If traditional SELEX method is used for aptamer identification, then aptamers can be selected with good binding properties, but the process requires multiple rounds (6-20 rounds) and extensive follow-up work including cloning and random selection, leading to high time consumption and labor intensity
Solution Approach 1:
The invention divides the aptamer identification process into distinct segments: (1) binding selection step to enrich aptamers, (2) spatially separated amplification to clone and sequence all enriched oligonucleotides individually, and (3) systematic assessment of binding properties. This segmentation eliminates the need for multiple SELEX rounds and random selection, allowing complete characterization of all bound oligonucleotides in parallel, thereby dramatically reducing time and labor while maintaining assessment precision.
Solution Approach 2:
The invention performs preliminary spatially separated amplification and sequencing of all oligonucleotides that bound in the selection step before conducting binding property assessments. This preliminary action ensures that all potential aptamers are identified and characterized systematically, eliminating the need for subsequent random selection and multiple selection rounds, thus reducing overall process time while improving completeness of identification.
2Reliability
If traditional SELEX method is used, then aptamers can be selected, but unfavorable selection conditions during any of the multiple rounds can cause loss of good binders
Solution Approach 1:
The invention performs spatially separated amplification and sequencing of all oligonucleotides that bound in the selection step as a preliminary action before any assessment or further selection. This ensures that all potential aptamers are captured and identified systematically in a single selection event, eliminating the risk of losing good binders due to unfavorable conditions in subsequent rounds while maintaining high identification efficiency.
Solution Approach 2:
The invention implements a feedback mechanism where all oligonucleotides bound in the selection step are amplified, sequenced, and their binding properties are assessed systematically. This complete feedback loop ensures that no potential aptamer is lost, as all bound oligonucleotides are characterized and evaluated, improving reliability without sacrificing productivity.
3Measurement precision
If traditional SELEX method is used with random selection of aptamers for further characterization, then some aptamers can be identified, but individual good binders can be lost because they are not randomly selected and therefore not recorded
Solution Approach 1:
The invention segments the characterization process by performing spatially separated amplification that creates individual amplificates for each oligonucleotide, followed by systematic sequencing and binding property assessment of all enriched oligonucleotides. This segmentation ensures complete characterization of all potential aptamers without relying on random selection, improving completeness while the automated nature of the process manages complexity.
Solution Approach 2:
The spatially separated amplification and sequencing process serves itself by automatically generating individual records for each oligonucleotide that bound in the selection step. This self-service mechanism ensures that all aptamers are systematically identified and characterized without requiring random human selection, improving completeness of characterization while the standardized process handles the complexity.
4Measurement precision
If multiple rounds of SELEX are performed to accumulate high affinity and specificity binders, then good binders can be obtained, but the process becomes extremely time-consuming and labor-intensive
Solution Approach 1:
The invention performs preliminary spatially separated amplification and sequencing of all oligonucleotides bound in a single selection step, capturing all potential high-affinity binders in one event. This preliminary action eliminates the need for multiple enrichment rounds, allowing systematic identification and assessment of all aptamers with high binding affinity and specificity, thereby maintaining measurement precision while dramatically improving development throughput.
Solution Approach 2:
The invention segments the aptamer development process into a single binding selection step followed by comprehensive spatially separated amplification, sequencing, and assessment of all enriched oligonucleotides. This segmentation allows complete characterization of all potential aptamers in parallel, achieving the same binding affinity and specificity as multiple rounds but with much higher productivity and reduced time consumption.
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 method significantly reduces the time and effort required for aptamer identification, increases yield, and provides better assessment of aptamer properties, enabling parallel sequencing and analysis of a large number of oligonucleotides, and offers higher specificity compared to traditional SELEX processes.
Implementation Method 1
bringing a mixture of oligonucleotides into contact with an aptamer target structure, and binding at least part of the oligonucleotides to the target structure
Implementation Method 2
the spatially separate amplification of all oligonucleotides that have bound to the aptamer target structure
Data Source
Figure 1
Figure 2A~2D
Figure 3
AI summary
The invention relates to a method for identifying aptamers, having the following steps - bringing a mixture of oligonucleotides into contact with an aptamer target structure and binding at least some of the oligonucleotides to the target structure, - separating the oligonucleotides which have been bound to the aptamer target structure from the aptamer target structure and from oligonucleotides that are not bound to the aptamer target structure, - amplifying individual oligonucleotides which were bound to the aptamer target structure in a physically separate manner and producing a plurality of physically separate amplicons, each amplicon predominantly containing one type of oligonucleotides, - specifying a specific marker for a plurality of the physically separate amplicons such that each of the marked amplicons can be uniquely identified using the specified marker of the amplicon, - sequencing oligonucleotides in a plurality of marked amplicons and assigning the marker that is specific for the amplicon to the sequence of the type of oligonucleotides in the amplicon for each amplicon examined by means of the sequencing process - analyzing the binding properties of the types of oligonucleotides to the aptamer target structure and assigning the analyzed binding properties to the specific markers of the amplicons and to the sequences of the types of oligonucleotides.