MITIGATION OF Cot-1 DNA DISTORTION IN NUCLEIC ACID HYBRIDIZATION

Abstract

A novel method of suppressing non-specific cross-hybridization between repetitive elements present in nucleic acid probes and corresponding repetitive elements in the target nucleic acid by using DNA synthesized to contain a plurality of repetitive elements while avoiding low and single copy sequences.

1 - 11 . (canceled) 
     
     
         12 . A method of synthesizing suppression nucleic acid comprising the steps of:
 identifying repetitive sequences in a representative genomic region; and   synthesizing said suppression nucleic acid by synthesizing nucleic acid sequences hybridizable with said identified repetitive sequences but not hybridizable with low copy sequences near or within said representative genomic region.   
     
     
         13 . The method of  claim 12 , said synthesized suppression nucleic acid being substantially free of low copy sequences. 
     
     
         14 . The method of  claim 12 , further comprising the step of selecting certain identified repetitive sequences for synthesis as said suppression nucleic acid based on their proximity to a low copy sequence of interest. 
     
     
         15 . The method of  claim 14 , said certain identified repetitive sequences being closest in proximity to said low copy sequence of interest. 
     
     
         16 . The method of  claim 14 , said synthesized suppression nucleic acid being free of sequences that are hybridizable with said low copy sequence of interest. 
     
     
         17 . The method of  claim 12 , further comprising the step of using said synthesized suppressive nucleic acid in a hybridization assay. 
     
     
         18 . The method of  claim 17 , said hybridization assay being selected from the group consisting of fluorescence in situ hybridization assays, microarray assays, and microsphere hybridization assays. 
     
     
         19 - 23 . (canceled) 
     
     
         24 . A method of accurately quantitating nucleic acid sequence copy numbers, said method comprising the steps of:
 preparing a first, spectrally-encoded, fluorescent microsphere having a first spectral address, and a second, spectrally-encoded, fluorescent microsphere having a second spectral address;   identifying a target genomic nucleic acid probe sequence by ascertaining the nucleotide-by-nucleotide sequence of a target nucleic acid sequence wherein the sequence of interest is suspected to reside;   synthesizing a low copy target probe derived from said identified target genomic nucleic acid probe sequence, said target probe comprising at least one low copy element and being substantially devoid of repetitive elements;   conjugating said target probe to said first microsphere;   synthesizing a reference probe selected to hybridize to a reference nucleic acid sequence of said target nucleic acid sequence;   conjugating said reference probe to a microsphere having a second spectral address;   identifying repetitive sequences in a representative genomic region;   synthesizing suppressive nucleic acid, said suppressive nucleic acid comprising sequences of sufficient homology to hybridize to said identified repetitive sequences, said suppressive nucleic acid substantially comprising repetitive elements and being substantially devoid of low copy elements;   reacting said suppressive nucleic acid with a chromosomal target sequence, thereby causing repetitive elements in said suppressive nucleic acid to hybridize with homologous repetitive elements in said chromosomal target sequence;   reacting said target probe to said chromosomal target sequence thereby causing low copy elements in said target probe to hybridize to homologous low copy elements in said chromosomal target sequence;   reacting said suppressive nucleic acid with a chromosomal reference sequence containing said chromosomal target sequence thereby causing repetitive elements in said suppressive nucleic acid to hybridize to homologous repetitive elements in said chromosomal reference sequence;   reacting said reference probe with said chromosomal reference sequence thereby causing said reference probe to hybridize to said chromosomal reference sequence;   detecting the hybridized target probe via said first spectral address;   detecting the hybridized reference probe via said second spectral address; and   quantifying the detected target probe by comparing the response of the detected hybridized target probe with the response of the detected hybridized reference probe.   
     
     
         25 . The method of  claim 24  wherein said suppressive DNA comprises repetitive elements, said repetitive elements corresponding to genomic repetitive elements adjacent to low copy elements in said target.