Cleavable Oligonucleotide Arrays for Synthesis Error Reduction
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Solution Overview
Problem
Existing methods of synthesizing polynucleotides, such as oligonucleotide arrays, face challenges in reducing error sequences, particularly at the edges of features due to misalignment and variability in droplet placement during the synthesis process, leading to impurities and defects in the final product.
Innovation Solution
The method involves covalently binding a cleavable linker moiety to a substrate surface, defining an interior area, and binding nucleotide moieties both via and outside of this linker, allowing for precise synthesis and reducing errors by distinguishing between oligonucleotides bound through the cleavable linker and those bound otherwise, thereby minimizing edge errors.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If in situ synthesis methods are used to fabricate oligonucleotide arrays, then large numbers of oligonucleotides can be synthesized efficiently, but error sequences occur at edge areas due to misalignment and variability in droplet placement
Solution Approach 1:
The feature area is segmented into interior and edge areas, with different handling strategies. Oligonucleotides in the interior area are synthesized with higher accuracy while edge area oligonucleotides are identified and removed, thus resolving the contradiction between maintaining high throughput and improving sequence accuracy.
Solution Approach 2:
Different quality standards are applied to different regions. The interior area oligonucleotides are designed to be the primary product with high sequence accuracy, while edge area oligonucleotides are accepted as lower quality and removed. This local differentiation allows high throughput synthesis while ensuring high accuracy for the intended product.
2Ease of manufacture
If droplet placement is performed during synthesis iterations, then oligonucleotide chains are built sequentially, but misalignment causes edge errors and reduces product purity
Solution Approach 1:
The problematic edge area oligonucleotides are extracted and removed from the final product. By identifying oligonucleotides that are not properly bound to the substrate (those in the edge area) and removing them, the method maintains ease of manufacture while significantly improving sequence fidelity of the remaining product.
Solution Approach 2:
The substrate surface is prepared with specific chemical properties beforehand to enable selective binding. The substrate is functionalized to preferentially bind oligonucleotides in the interior area while leaving edge area oligonucleotides unbound, allowing preliminary separation before final product collection and thus improving sequence fidelity.
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 significantly reduces the incidence of failure sequences in oligonucleotide synthesis, with oligonucleotides bound via the cleavable linker moiety exhibiting lower error rates compared to those bound at the edge areas, enhancing the purity and accuracy of the synthesized oligonucleotides.
Implementation Method 1
covalently binding a cleavable linker moiety to the surface
Data Source
AI summary
Oligonucleotide arrays having features that include cleavable oligonucleotides are disclosed, as well as methods of making such arrays. Methods of synthesizing an oligonucleotide on a surface of a substrate are described.