Method for the discovery of high-affinity, high specificity oligonucleotide and derivatized oligonucleotide sequences for target recognition

摘要

A method is disclosed to obtain oligonucleotide sequences with high affinity to target molecules. By design, the oligonucleotides have a defined primary and secondary structure. The affinity for binding to target species is classified or quantified by assay measurements using physical measurements rather than being based primarily on separations. Targets include but are not limited to proteins, polymers, biological membranes including cells and organelles and small molecules.

主权项

1. A method of obtaining combimers that bind to a target at a desired binding affinity, comprising the steps (a) through (d) in the following order:
(a) designing an oligonucleotide library comprising oligonucleotides or analogs thereof having known primary and secondary structure, each of which comprises a constant region and a variable region comprising N enumerated base positions in a combimer loop region of the oligonucleotide or analog thereof, wherein each enumerated base position comprises one of X selected nucleotide bases, such that the oligonucleotide library comprises up to XN different oligonucleotides or analogs thereof, and wherein the nucleotide sequence is systematically modified to create a family of sequence variants, and wherein the region of the nucleotide sequence which is varied is said to be enumerated; (b) synthesizing at least some of the oligonucleotides or analogs thereof of the oligonucleotide library; (c) classifying or quantifying the binding affinity of the combimer loop region of each oligonucleotide or analog thereof for the target; and (d) identifying combimers with the desired binding affinity from the library of oligonucleotides or analogs thereof,wherein the combimer of step (d) is a member of the oligonucleotide library of (a) which comprises the known primary and secondary structure of step (a) and wherein N enumerated base positions comprises 2-40 nucleotides, and wherein the oligonucleotide library of (a) is reusable with multiple targets.