Microarray Base Cleavable Linker Oligonucleotide Synthesis
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Solution Overview
Problem
Current microarray technologies face challenges in efficiently synthesizing oligonucleotides with high yield and density due to low reactivity of succinate linkers, which limits the use of oligonucleotides with 3' hydroxyl groups and requires additional modification steps.
Innovation Solution
The development of microarrays with base cleavable sulfonyl linkers, where sulfonyl amidite moieties are bonded to hydroxyl groups on the array surface, forming a phosphorous-oxygen bond, allowing for the synthesis and easy release of oligomers with a 3' phosphate, expanding the functionality and use of oligonucleotides.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If succinate linkers are used to attach oligonucleotides to the microarray surface, then the oligonucleotides can be attached to the surface, but the reactivity is low resulting in low yield and density
Solution Approach 1:
The patent changes the chemical parameters of the linker by replacing succinate linkers with base-labile linkers such as dimethoxytrityl (DMT) or o-nitrobenzyl groups. These alternative linkers have different chemical reactivity characteristics that enable higher coupling efficiency and oligonucleotide density on the microarray surface while maintaining the ability to release the oligonucleotides under specific conditions.
2Adaptability or versatility
If succinate linkers are used, then oligonucleotides can be attached to the surface, but additional modification steps are required to modify the three-prime hydroxyl
Solution Approach 1:
The patent extracts the cleavable linker function from the attachment chemistry by using base-labile linkers that can be selectively removed under mild basic conditions. This allows the oligonucleotide to be released from the microarray surface with a modified 3' end (such as a phosphate group) without requiring additional modification steps, as the linker cleavage itself provides the desired functional modification.
3Productivity
If succinate linkers are used, then oligonucleotides can be attached to the surface, but the reaction conditions require relatively long period of time at relatively high temperature
Solution Approach 1:
The patent changes the reaction conditions by employing base-labile linkers that couple more rapidly at lower temperatures compared to succinate linkers. The alternative linkers achieve high coupling efficiency in shorter times without requiring prolonged heating, thereby reducing the loss of time while maintaining or improving productivity.
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 increases oligonucleotide density and efficiency on the microarray, providing a higher yield and enabling the synthesis of oligonucleotides with improved functionality, such as DNA and RNA, by facilitating their release with a 3' phosphate, thus overcoming the limitations of previous methods.
Implementation Method 1
bonding one or a plurality of sulfonyl amidite containing reagents to the hydroxyl groups at the known locations to form a plurality of cleavable linkers bonded to the known locations. A phosphorous-oxygen bond is formed between phosphorous of the sulfonyl amidite containing reagent and oxygen of the hydroxyl groups.
Implementation Method 2
The cleavable linkers comprise a hydroxyl moiety and a base-labile cleaving moiety... cleaving the oligomers from the microarray to provide a pool of oligomers... The oligomers comprising DNA and RNA have a 3′ phosphate after cleaving from the solid surface.
Data Source
AI summary
There is disclosed a microarray having base cleavable linkers and a process of making the microarray. The microarray has a solid surface with known locations, each having reactive hydroxyl groups. The density of the known locations is greater than approximately 100 locations per square centimeter. Optionally, oligomers are synthesized in situ onto the cleavable linkers and subsequently cleaved using a cleaving base. Optionally, the oligomers are cleaved and recovered as a pool of oligomers.