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US 9,040,678 B2Granted
Functionalization and purification of molecules by reversible group exchange
AssigneeIllumina, Inc.
InventorsFrank J. Steemers, Kevin L. Gunderson, Kerri York, Ryan Christopher Smith
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Abstract
Embodiments of the present disclosure include methods and compositions for functionalizing molecules, such as oligonucleotides, with functional groups, including polyhistidine tags useful in affinity methods. Some embodiments include methods for modifying and purifying complex mixtures of molecules by exchange of functional tags.
Claims20
A method for modifying a nucleic acid, said method comprising:
- (a) performing a first exchange reaction comprising contacting:
- a nucleic acid comprising an aldehyde group, and
- a bifunctional linking reagent comprising a hydrazine or amine moiety under conditions to form an imine bond coupling the nucleic acid to the bifunctional linking reagent;
- wherein the bifunctional linking reagent further comprises a functional tag;
- (b) contacting the sample with a first binding partner to the first functional tag;
- and
- (c) performing a first purification step comprising separating the first binding partner from the sample, thereby isolating a nucleic acid from a sample;
- (d) performing a second exchange reaction comprising contacting the nucleic acid isolated in step (c) with an exchange reagent under conditions sufficient for the imine bond to be broken between the nucleic acid and the bifunctional linking reagent and for a new imine bond to form between the exchange reagent and the nucleic acid;
- (f) contacting the nucleic acid with a second binding partner to the second functional tag;
- and
- (g) separating the second binding partner from the nucleic acid that is not associated with the second affinity tag, thereby isolating the nucleic acid.
8 more elements
Dependencies are inferred from the claim text.