Competitive probes for engineering signal generation

Abstract

A method of identifying a first target nucleic acid comprising, providing a sample comprising the first target nucleic acid, providing a first set of paired oligonucleotides with complementarity to the first target nucleic acid, the first set of paired oligonucleotides comprising a first ratio of (a) first competitive oligonucleotides to (b) first signal oligonucleotides comprising a signal tag, wherein the competitive oligonucleotides compete with the signal oligonucleotides for binding to the first target nucleic acid, amplifying the first target nucleic acid with the polymerase chain reaction, thereby degrading the first signal oligonucleotide and permitting generation of a first signal, generating the first signal, measuring intensity of the first signal, and correlating the intensity of the first signal to the first ratio, thereby identifying the first target nucleic acid.

What is claimed is: 
     
         1 . Method of identifying a first target nucleic acid comprising,
 providing a sample comprising the first target nucleic acid,   providing a first set of paired oligonucleotides with complementarity to the first target nucleic acid, said first set of paired oligonucleotides comprising a first ratio of (a) first competitive oligonucleotides to (b) first signal oligonucleotides comprising a first signal tag,   wherein the first competitive oligonucleotides compete with the first signal oligonucleotides for binding to the first target nucleic acid,   amplifying the first target nucleic acid with the polymerase chain reaction, thereby degrading the first signal oligonucleotide and permitting generation of a first signal,   generating the first signal,   measuring intensity of the first signal, and   correlating the intensity of the first signal to the first ratio, thereby identifying the first target nucleic acid.   
     
     
         2 . The method of  claim 1 , wherein the sample comprises a second target nucleic acid. 
     
     
         3 . The method of  claim 2 , further comprising identifying the second target nucleic acid by:
 providing a second set of paired oligonucleotides with complementarity to the second target nucleic acid, said second set of paired oligonucleotides comprising a second ratio of (a) second competitive oligonucleotides to (b) second signal oligonucleotides comprising a second signal tag,   wherein the second competitive oligonucleotides compete with the second signal oligonucleotides for binding to the second target nucleic acid,   amplifying the second target sequence with the polymerase chain reaction,   thereby degrading the second signal oligonucleotide and permitting generation of a second signal,   generating the second signal,   measuring intensity of the second signal, and   correlating the intensity of the second signal to the second ratio, thereby identifying the second target nucleic acid.   
     
     
         4 . The method of  claim 3 , wherein the first signal oligonucleotides and the second signal oligonucleotides comprise the same signal tag. 
     
     
         5 . The method of  claim 3 , wherein the first target nucleic acid and the second target nucleic acid are identified from differing signal intensities generated during the same polymerase chain reaction. 
     
     
         6 . A composition comprising:
 a first set of paired oligonucleotides with complementarity to a first target nucleic acid,   said first set of paired oligonucleotides comprising a first ratio of (a) competitive oligonucleotides to (b) signal oligonucleotides comprising a signal tag,   wherein the competitive oligonucleotides are configured to compete with the signal oligonucleotides for binding to the first target nucleic acid during amplification of the first target nucleic acid with the polymerase chain reaction,   wherein the signal oligonucleotides are adapted to generate a first signal having a first intensity when degraded by a nucleic acid polymerase during the polymerase chain reaction and excited by an energy source,   and wherein the first intensity correlates with the first ratio.   
     
     
         7 . Method of identifying a first target molecule comprising,
 providing a sample comprising the first target molecule,   providing a first set of paired capture molecules that bind to the first target molecule, said first set of paired capture molecules comprising a first ratio of (a) first competitive capture molecules to (b) first signal capture molecules comprising a signal tag,   wherein the competitive capture molecules compete with the signal capture molecules for binding to the first target molecule,   generating the first signal,   measuring intensity of the first signal, and   correlating the intensity of the first signal to the first ratio, thereby identifying the first target molecule.   
     
     
         8 . The method of  claim 7 , wherein the sample comprises a second target molecule. 
     
     
         9 . The method of  claim 8 , further comprising identifying the second target molecule by:
 providing a second set of paired capture molecules that binds to the second target molecule, said second set of paired capture molecules comprising a second ratio of (a) second competitive capture molecules to (b) second signal capture molecules comprising the signal tag,   wherein the second competitive capture molecules compete with the second signal capture molecules for binding to the second target molecule,   generating the second signal,   measuring intensity of the second signal, and   correlating the intensity of the second signal to the second ratio, thereby identifying the second target molecule.   
     
     
         10 . The method of  claim 9 , wherein the first signal capture molecules and the second signal capture molecules comprise the same signal tag. 
     
     
         11 . The method of  claim 8 , wherein the first target capture molecules and the second target capture molecules are identified from differing signal intensities generated. 
     
     
         12 . A composition comprising:
 a first set of paired capture molecules with complementarity to a first target molecule,   said first set of paired capture molecules comprising a first ratio of (a) competitive capture molecules to (b) signal capture molecules comprising a signal tag,   wherein the competitive capture molecules are configured to compete with the signal capture molecules for binding to the target molecule,   wherein the signal capture molecules are adapted to generate a first signal having a first intensity when excited by an energy source,   and wherein the first intensity correlates with the first ratio.   
     
     
         13 . The composition of  claim 12 , wherein the target molecule comprises a protein or a functional fragment thereof. 
     
     
         14 . The composition of  claim 12 , wherein the target molecule comprises a antigen. 
     
     
         15 . The composition of  claim 12 , wherein the target molecule comprises a aptamer. 
     
     
         16 . The composition of  claim 12 , wherein the target molecule comprises a nucleic acid. 
     
     
         17 . The composition of  claim 12 , wherein the capture molecule comprises a protein or a functional fragment thereof. 
     
     
         18 . The composition of  claim 12 , wherein the capture molecule comprises an antibody or a functional fragment thereof. 
     
     
         19 . The composition of  claim 12 , wherein the capture molecule comprises an aptamer. 
     
     
         20 . Method of altering amplification level of a target nucleic acid comprising,
 providing a sample comprising the target nucleic acid,   providing a set of paired oligonucleotides with complementarity to the target nucleic acid, said set of paired oligonucleotides comprising a ratio of (a) competitive oligonucleotides to (b) functional oligonucleotides,   wherein the competitive oligonucleotides compete with the functional oligonucleotides for binding to the target nucleic acid and are not extendible by a nucleic acid polymerase, wherein the functional oligonucleotides are extendible by a nucleic acid polymerase as primers for amplifying the target nucleic acid by the polymerase chain reaction (PCR),   and   amplifying the target nucleic acid with PCR.   
     
     
         21 . The method of  claim 20 , wherein the competitive oligonucleotides comprise at least one modified end. 
     
     
         22 . The method of  claim 21 , wherein the modified end prevents enzymatic extension during amplification. 
     
     
         23 . The method of  claim 22 , wherein the modified end comprises a 3′ end lacking a hydroxyl group. 
     
     
         24 . The method of  claim 22 , wherein the modified end comprises a 5′ end lacking a phosphate group. 
     
     
         25 . The method of  claim 22 , wherein the modified end comprises a 5′ capped end. 
     
     
         26 . The method of  claim 22 , wherein the modified end comprises a 3′ capped end. 
     
     
         27 . The method of  claim 20 , wherein a total number of molecules in the reaction remains constant. 
     
     
         28 . A composition comprising:
 a set of paired oligonucleotides with complementarity to a target nucleic acid,   said set of paired oligonucleotides comprising a ratio of (a) competitive oligonucleotides to (b) functional oligonucleotides,   wherein the competitive oligonucleotides are configured to compete with the functional oligonucleotides for binding to the nucleic acid target during amplification of the first target nucleic acid with the polymerase chain reaction and are not extendible by a nucleic acid polymerase.