Methods of generating nanoarrays and microarrays
Inventors
Gremyachinskiy, Dmitriy • GALIMIDI, Rachel • Mallick, Parag • Patel, Sujal M.
Assignees
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Publication Number
US-11603383-B2
Publication Date
2023-03-14
Expiration Date
Abstract
The methods described herein provide a means of producing an array of spatially separated proteins. The method relies on covalently attaching each protein of the plurality of proteins to a structured nucleic acid particle (SNAP), and attaching the SNAPs to a solid support.
Core Innovation
The invention provides heterobifunctional crosslinker chemistry and spacer/cross-bridge considerations for bioconjugation, including intermediate forms that differ in reactivity. The approach includes crosslinkers with functional reactive groups suitable for attaching biomolecules to solid supports, with examples such as NHS ester–maleimide chemistry, cleavable cross-bridges, and PEG spacers that provide defined spacing.
The invention further provides extensive crosslinker examples and functional reactive groups for attaching biomolecules to solid supports, including azide/alkyne and other bioorthogonal or surface-reactive chemistries. The document includes examples of SNAP coupling to surfaces, including approaches described as DNA SNAP and SNAPs configured for conjugation via click chemistry.
The document outlines the production, purification, and characterization of nucleic-acid “DNA SNAPs,” followed by conjugation assays using solid supports such as chips/arrays and particles. It describes characterization of size/occupancy/brightness and conjugation using click conjugation (DBCO–azide) with fluorescence-based detection and specificity controls.
Claims Coverage
The partial content provided includes one independent claim (clm-00001) directed to attaching a protein to a solid support using seed oligonucleotide polymerization into a structured nucleic acid particle. The independent claim includes three main inventive features.
Seed oligonucleotide covalently attached to a protein
A seed oligonucleotide is provided, wherein the seed oligonucleotide is covalently attached to a protein.
Hybridization to an immobilized self-hybridizing oligonucleotide on a solid support
The seed oligonucleotide is hybridized to an immobilized oligonucleotide, wherein the immobilized oligonucleotide is covalently attached to a solid support and has a region of internal complementarity that is self-hybridizing.
Polymerization to form a structured nucleic acid particle that attaches the protein to the solid support
The seed oligonucleotide is grown by polymerization, thereby producing a structured nucleic acid particle comprising a self-hybridized region of internal complementarity, wherein the protein is attached to the solid support via the structured nucleic acid particle.
Overall, the claim coverage centers on connecting a protein to a solid support through an intermediate nucleic-acid hybridization and polymerization step that produces a structured nucleic acid particle with internal complementarity.
Stated Advantages
Not explicitly described in patent.
Documented Applications
Not explicitly described in patent.
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