Selex and photoSELEX

A method for identifying a photoaptamer, the method comprising:

• a) preparing a candidate mixture of nucleic acids comprising at least one non-photoreactive placeholding modified pyrimidine;
• b) contacting the candidate mixture with a target, wherein nucleic acids having an increased affinity to the target relative to the candidate mixture may be partitioned from the remainder of the candidate mixture;
• c) partitioning the increased affinity nucleic acids from the remainder of the candidate mixture;
• d) amplifying the increased affinity nucleic acids to yield a nucleic acid ligand-enriched mixture of nucleic acids;
• e) repeating (b)-(d) as desired;
• f) producing from said nucleic acid ligand-enriched mixture of nucleic acids a candidate photoaptamer or a mixture of candidate photoaptamers by replacing in each nucleic acid of the ligand-enriched mixture of nucleic acids one or more non-photoreactive placeholding pyrimidines with a photoreactive pyrimidine;
• g) contacting said candidate photoaptamer(s) with said target wherein a candidate photoaptamer-target complex is formed;
• h) irradiating said candidate photoaptamer-target complex;
• i) determining whether said candidate photoaptamer-target complex has photocrosslinked;
• j) repeating (f)-(i) as desired;
• and
• k) identifying at least one photoaptamer to the target.

A method for identifying an aptamer, the method comprising:

• a) preparing a candidate mixture of nucleic acids;
• b) contacting the candidate mixture with a biotinylated target molecule, wherein nucleic acids having an increased affinity to the target molecule relative to other nucleic acids in the candidate mixture bind the target molecule, forming nucleic acid-target molecule complexes;
• c) partitioning the nucleic acid-target molecule complexes from the candidate mixture by contacting the candidate mixture with a solid support having streptavidin bound to the solid support and washing the solid support with a solution containing biotin;
• d) dissociating the nucleic acid-target molecule complexes to form free nucleic acids;
• e) amplifying the free nucleic acids to yield a mixture of nucleic acids enriched with nucleic acids that are capable of binding to the target molecule with increased affinity;
• f) repeating (b) through (e) as desired;
• and
• g) identifying at least one aptamer to the target molecule.