Patent ID: 6022691
Filing Date: 2000-02-08
Classification: A61K,C12Q

Abstract:
A method of specifically detecting a chemical or drug in a sample comprising contacting the sample with an oligonucleotide having specific activity for a target biomolecule, wherein said oligonucleotide is identified by the following steps:(a) preparing a set of randomized oligonucleotides;(b) subfractionating the set of randomized oligonucleotides to provide subfractions of said oligonucleotides;(c) assaying the subfractions of randomized oligonucleotides for activity against a target biomolecule;(d) selecting the subfraction having the highest activity;(e) assaying the oligonucleotides of the selected subfraction for activity against a target biomolecule;(f) separating active from inactive oligonucleotides;(g) recovering said active oligonucleotides;(h) extending a polyA on said active oligonucleotide in a 3' direction to form a first strand;(i) hybridizing said first strand to a first chimeric primer having a 5' known sequence and a 3' polydT portion;(j) forming a cDNA strand, complementary to the active oligonucleotide, using the first chimeric primer;(k) extending said cDNA formed in (j) in a 3' direction by the addition of polyA;(l) separating said oligonucleotide and cDNA strand;(m) hybridizing the polyA portion of said cDNA strand to a second chimeric primer having a 5' known sequence and a 3' polydT portion;(n) filling in the recessed 3' ends to form two complementary strands;(o) separating the strands;(p) amplifying the strands formed in (m) using the first and second chimeric primers by polymerase chain reaction;(q) excising active oligonuclcotide from flanking regions;(r) recovering active oligonucleotide; and(s) performing steps (b) through (r) iteratively to determine an active oligonucleotide; anddetecting the presence or absence of binding, whereby the presence of binding is indicative of the presence of the chemical or drug in said sample.