Oligonucleotide Codon Decoding via Primer Extension
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Solution Overview
Problem
Current methods for obtaining structural information about encoded molecules, particularly those formed by reactions of multiple chemical entities, are inefficient and require complex decoding processes, often involving multiple steps and host organisms like E. coli for PCR and sequencing.
Innovation Solution
A method involving mixing a primer oligonucleotide with an identifier oligonucleotide containing codons that encode the chemical entities involved in the molecule's formation, followed by an extension reaction using polymerase and (deoxy)ribonucleotide triphosphates, allowing for the evaluation of codon presence, absence, or relative abundance to deduce the molecule's structure.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If traditional PCR and sequencing methods are used to decode identifier oligonucleotides, then structural information about encoded molecules can be obtained, but the process becomes complex and time-consuming
Solution Approach 1:
The invention extracts and analyzes only the relevant codon regions from the identifier oligonucleotide using specific primers, rather than performing full PCR amplification and sequencing. This extracts the essential structural information while eliminating unnecessary complex steps
Solution Approach 2:
The method creates simplified copies of specific codon regions through primer extension reactions, generating readable signals that represent the encoded molecular structure without requiring full sequencing of the identifier oligonucleotide
2Loss of information
If multiple decoding steps are performed to identify chemical entities in encoded molecules, then complete structural information is obtained, but the time required for analysis increases
Solution Approach 1:
The identifier oligonucleotide is segmented into discrete codon regions, each representing a specific chemical entity. Primers are designed to target specific codon positions, allowing parallel analysis of multiple segments in a single reaction setup
Solution Approach 2:
The primers are pre-designed with specific sequences that complement target codons. The extension reaction conditions are pre-optimized to selectively amplify only present codons, eliminating the need for multiple sequential decoding steps
3Reliability
If host organisms like E. coli are used for PCR and sequencing, then reliable structural data is obtained, but the ease of operation decreases
Solution Approach 1:
The invention replaces the biological system (host organisms like E. coli) with a biochemical system (in vitro extension reactions). This substitution maintains reliability through controlled enzymatic reactions while dramatically improving ease of operation by eliminating microbial culture and handling requirements
4Loss of information
If comprehensive decoding of all codons is performed, then complete molecule structure is determined, but the productivity of the screening process decreases
Solution Approach 1:
The method performs partial decoding by analyzing only the specific codon regions relevant to the screening objective. By using targeted primers that extend only from present codons, the system obtains sufficient structural information without the overhead of comprehensive full-length sequencing
Solution Approach 2:
The extension reactions produce detectable signals (analogous to color changes) that directly indicate the presence or absence of specific codons. This provides immediate readable output that can be rapidly processed, maintaining productivity while determining complete molecular structure
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 provides rapid and reliable structural information about encoded molecules, enabling the identification of successful chemical entities and facilitating the formation of second-generation libraries with enhanced affinity for targets, thus streamlining the drug discovery process.
Implementation Method 1
the presence, absence or relative abundance of a codon is evaluated by mixing a primer with the identifier oligonucleotide in the presence of a polymerase and substrate (deoxy)ribonucleotide triphosphates and measuring the extension reaction
Data Source
AI summary
In one aspect, the present invention relates to a method for obtaining structural information about an encoded molecule. The encoded molecule may be produced by a reaction of a plurality of chemical entities and may be capable of being connected to an identifier oligonucleotide containing codons informative of the identity of the chemical entities which have participated in the formation of the encoded molecule. In a certain embodiment, primers are designed complementary to the codons appearing on the identifier oligonucleotide, and the presence, absence or relative abundance of a codon is evaluated by mixing a primer with the identifier oligonucleotide in the presence of a polymerase and substrate (deoxy)ribonucleotide triphosphates and measuring the extension reaction. In another aspect, the invention provides a method for selecting compounds which binds to a target. More specifically, the invention relates to a method in which a target associated with an oligonucleotide initially is mixed with a library of complexes, each complex comprising a display molecule and an oligonucleotide identifying said display molecule. Next, due an increased proximity, the target oligonucleotide is coupled to the identifier oligonucleotide of complexes having a display mole-cute with affinity towards the target. In a final stage the coupled nucleotides are analysed to deduce at least the identity of the display molecule.