Encoding Oligonucleotide Tags for High-Complexity Library Synthesis
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Solution Overview
Problem
Current methods for producing combinatorial libraries face challenges in synthesizing libraries of sufficient complexity and identifying active molecules, as increasing complexity reduces the concentration of individual library members, making it difficult to identify active compounds in high-complexity libraries.
Innovation Solution
A method utilizing a 'split and pool' strategy to synthesize libraries with encoding oligonucleotide tags, where a solution comprising an initiator linked to an oligonucleotide is divided, reacted with unique building blocks, and then pooled, allowing for the synthesis of vast numbers of distinct molecules with each member encoded by a unique oligonucleotide sequence.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If the number of distinct members within the library is increased to raise complexity, then the probability of containing active molecules is improved, but the concentration of any particular library member is lowered making identification difficult
Solution Approach 1:
The library synthesis process is segmented into multiple cycles of splitting and pooling. In each cycle, the library is divided into separate reaction vessels where different building blocks are added, then pooled together. This segmentation allows for the systematic construction of high-complexity libraries while maintaining track of individual members through encoding tags.
Solution Approach 2:
Encoding oligonucleotide tags are copied and attached to each library member during synthesis. These tags serve as molecular copies or identifiers that allow researchers to determine the structure of active compounds by sequencing the tags, rather than needing to physically isolate and analyze the actual compound molecules.
2Reliability
If DNA encoding methods are used to identify library members, then amplifiable tags can be obtained, but the production of very large libraries has not been successfully demonstrated yet
Solution Approach 1:
The split and pool methodology enables continuous library synthesis through multiple sequential cycles. Each cycle builds upon the previous one, continuously adding complexity and diversity to the library while maintaining the amplifiable DNA tag throughout the process, thereby achieving both reliability and high productivity.
Solution Approach 2:
The encoding strategy uses nested oligonucleotide tags that are incorporated at each synthesis cycle. The tags are built in a nested manner where each cycle adds to the existing tag structure, allowing the tags to scale with the library complexity while maintaining their amplifiable nature.
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 synthesis of large libraries with high-fidelity oligonucleotide tags, facilitating the identification of active compounds by amplifying and sequencing the tags to determine the structure of binding molecules, allowing for multiple rounds of selection and amplification.
Implementation Method 1
reacting the initial oligonucleotide with an incoming oligonucleotide which identifies the building block in the presence of an enzyme which catalyses ligation of the initial oligonucleotide and the incoming oligonucleotide
Data Source
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AI summary
The present invention provides a method of synthesizing libraries of molecules which include an encoding oligonucleotide tag, and identifying compounds which bind to said biological target.