Oligonucleotide Paints

Abstract

Novel methods for making high resolution oligonucleotide paints are provided. Novel, high resolution oligonucleotide paints are also provided.

1 . A method of making a set of high resolution oligonucleotide paints comprising:
 providing at least one solid support having a plurality of synthetic, single stranded oligonucleotide sequences attached thereto, wherein a portion of each of the plurality of synthetic, single stranded oligonucleotide sequences is complementary to a portion of a specific chromosome sequence;   synthesizing a plurality of complementary strands, each of which is complementary to a synthetic, single stranded oligonucleotide sequence attached to the at least one solid support;   removing the plurality of complementary strands from the at least one solid support;   amplifying the plurality of complementary strands; and   labelling the plurality of complementary strands to produce a set of high resolution oligonucleotide paints, wherein the set of oligonucleotides paints has a resolution of about two kilobases or fewer.   
     
     
         2 . The method of  claim 1 , wherein the set of oligonucleotides paints has a resolution of about one kilobase or fewer. 
     
     
         3 . The method of  claim 1 , wherein the set of oligonucleotides paints has a resolution of about 100 bases or fewer. 
     
     
         4 . The method of  claim 1 , wherein the set of oligonucleotides paints has a resolution of between about 20 bases and about 30 bases. 
     
     
         5 . The method of  claim 1 , wherein 14 bases at each of the 3′ and 5′ ends of the oligonucleotide sequences are primer sequences and 32 bases internal to the primer sequences are complementary to a specific chromosome sequence. 
     
     
         6 . The method of  claim 1 , wherein each of the oligonucleotide paints has a detectable label attached thereto. 
     
     
         7 . The method of  claim 1 , wherein each of the oligonucleotide paints has a retrievable label attached thereto. 
     
     
         8 . The method of  claim 7 , wherein the retrievable label further binds a moiety selected from the group consisting of a protein, a peptide, a DNA sequence, an RNA sequence and a carbohydrate. 
     
     
         9 . The method of  claim 8 , wherein the retrievable moiety is exposed to light, heat or a chemical to activate binding of the retrievable label to a moiety selected from the group consisting of a protein, a peptide, a DNA sequence, an RNA sequence and a carbohydrate. 
     
     
         10 . A method of making a set of oligonucleotide paints comprising:
 providing at least one solid support having a plurality of synthetic, single stranded oligonucleotide sequences attached thereto, wherein a portion of each of the plurality of synthetic, single stranded oligonucleotide sequences is complementary to a portion of a specific chromosome sequence, and wherein each specific chromosome sequence excludes highly repetitive elements;   synthesizing a plurality of complementary strands, each of which is complementary to a synthetic, single stranded oligonucleotide sequence attached to the at least one solid support;   removing the plurality of complementary strands from the at least one solid support;   amplifying the plurality of complementary strands; and   labelling the plurality of complementary strands to produce a set of oligonucleotide paints.   
     
     
         11 . The method of  claim 10 , wherein each specific chromosome sequence excludes repetitive elements present in the genome as two copies, three copies or four copies in a haploid genome. 
     
     
         12 . The method of  claim 10 , wherein the length of each of the oligonucleotide sequences is about 60 bases. 
     
     
         13 . The method of  claim 12 , wherein 14 bases at each of the 3′ and 5′ ends of an oligonucleotide sequence are primer sequences and 32 bases internal to the primer sequences are complementary to a chromosome sequence. 
     
     
         14 . The method of  claim 10 , wherein each of the oligonucleotide paints has a detectable label attached thereto. 
     
     
         15 . The method of  claim 10 , wherein each of the oligonucleotide paints has a retrievable label attached thereto. 
     
     
         16 . The method of  claim 15 , wherein the retrievable label further binds a moiety selected from the group consisting of a protein, a peptide, a DNA sequence, an RNA sequence and a carbohydrate. 
     
     
         17 . The method of  claim 15 , wherein the retrievable moiety is exposed to light, heat or a chemical to activate binding of the retrievable label to a moiety selected from the group consisting of a protein, a peptide, a DNA sequence, an RNA sequence and a carbohydrate. 
     
     
         18 . The method of  claim 14 , wherein the detectable label is a fluorescent label. 
     
     
         19 . The method of  claim 14 , wherein the set of oligonucleotide paints provides at least 24 spectrally resolvable labels. 
     
     
         20 . The method of  claim 14 , wherein the set of oligonucleotide paints provides one spectrally resolvable color. 
     
     
         21 . The method of  claim 14 , wherein the set of oligonucleotide paints provides a spectrally resolvable label for each chromosome of an organism. 
     
     
         22 . The method of  claim 14 , wherein the set of oligonucleotide paints provides a spectrally resolvable label for one or more sub-chromosomal regions of an organism. 
     
     
         23 . The method of  claim 14 , wherein the at least one solid support is at least one microarray. 
     
     
         24 . The method of  claim 23 , wherein at least 25 microarrays are provided. 
     
     
         25 . The method of  claim 23 , wherein at least 100 microarrays are provided. 
     
     
         26 . The method of  claim 10 , wherein the step of amplifying includes providing a plurality of primers each having a portion that is complementary to a portion of a complementary strand or a portion of a single stranded oligonucleotide sequence. 
     
     
         27 . The method of  claim 26 , wherein the primers are universal primers. 
     
     
         28 . The method of  claim 26 , wherein at least a portion of each of the primer sequences is removable after said amplification step. 
     
     
         29 . The method of  claim 27 , wherein the universal primers comprise between one and 1000 different sequences. 
     
     
         30 . The method of  claim 27 , wherein the universal primers comprise at least 1000 different sequences. 
     
     
         31 . A palette of oligonucleotide paints comprising:
 a plurality of oligonucleotide sequences, wherein each oligonucleotide sequence is complementary to a single type of mutation corresponding to one of a specific set of chromosome abnormalities associated with a disorder, and wherein the set comprises at least 50 different mutations.   
     
     
         32 . The palette of oligonucleotide paints of  claim 31 , wherein the set comprises at least 100 different types of mutations. 
     
     
         33 . The palette of oligonucleotide paints of  claim 31 , wherein the set comprises at least 1000 different types of mutations. 
     
     
         34 . A set of oligonucleotide paints produced by the method of  claim 1 . 
     
     
         35 . A method of detecting a chromosome rearrangement in a biological sample comprising:
 providing a biological sample;   contacting the biological sample with the set of oligonucleotide paints of  claim 34 ;   detecting binding of the set of oligonucleotide paints;   comparing the binding of the set of oligonucleotide paints to a standard; and   detecting a chromosome rearrangement if binding of the set of oligonucleotide paints differs from the standard.   
     
     
         36 . The method of  claim 35 , wherein the chromosome rearrangement is selected from the group consisting of translocation, insertion, inversion, deletion, duplication, transposition, aneuploidy, polyploidy, complex rearrangement and telomere loss. 
     
     
         37 . A method of making a set of oligonucleotide paints comprising:
 providing at least one solid support having a plurality of synthetic, single-stranded oligonucleotide sequences attached thereto, wherein a portion of each of the plurality of synthetic, single-stranded oligonucleotide sequences is complementary to a portion of a specific chromosome sequence;   removing the plurality of synthetic oligonucleotide sequences from the at least one solid support;   amplifying the plurality of synthetic oligonucleotide sequences to generate amplified, synthetic oligonucleotide sequences; and   labelling the amplified, synthetic oligonucleotide sequences to produce a set of oligonucleotide paints, wherein each oligonucleotide paint has a resolution of about two kilobases or fewer.   
     
     
         38 . The method of  claim 37 , wherein the set of oligonucleotide paints is fluorescently labelled. 
     
     
         39 . The method of  claim 37 , wherein the set of oligonucleotide paints provides at least 24 spectrally resolvable labels. 
     
     
         40 . The method of  claim 37 , wherein the set of oligonucleotide paints provides one spectrally resolvable color. 
     
     
         41 . The method of  claim 37 , wherein the set of oligonucleotide paints provides a spectrally resolvable label for each chromosome of an organism. 
     
     
         42 . The method of  claim 37 , wherein the set of oligonucleotide paints provides a spectrally resolvable label for one or more sub-chromosomal regions of an organism. 
     
     
         43 . The method of  claim 37 , wherein the at least one solid support is at least one microarray. 
     
     
         44 . The method of  claim 43 , wherein at least 25 microarrays are provided. 
     
     
         45 . The method of  claim 43 , wherein at least 100 microarrays are provided. 
     
     
         46 . The method of  claim 37 , wherein the step of amplifying includes providing a plurality of primers, each of which is complementary to a portion of a synthetic oligonucleotide sequence. 
     
     
         47 . The method of  claim 46 , wherein the primers are universal primers. 
     
     
         48 . The method of  claim 46 , wherein a portion of each of the primer sequences is removable after said amplification step. 
     
     
         49 . A set of oligonucleotide paints produced by the method of  claim 37 . 
     
     
         50 . An article of manufacture for making a set of high resolution oligonucleotide paints comprising:
 a plurality of microarrays, each microarray having a plurality of synthetic oligonucleotide sequences attached thereto, wherein a portion of each of the plurality of synthetic oligonucleotide sequences is complementary to a portion of a specific chromosome sequence, wherein the sum of synthetic oligonucleotide that are complementary corresponds to between about 5% and 25% of a genome of interest, and wherein the set of oligonucleotides paints has a resolution of about two kilobases or fewer.   
     
     
         51 . The article of manufacture of  claim 50 , wherein the plurality of synthetic oligonucleotide sequences is complementary to at least 25% of a genome. 
     
     
         52 . The article of manufacture of  claim 50 , further including a plurality of primers. 
     
     
         53 . The article of manufacture of  claim 52 , wherein the plurality of primers are universal primers. 
     
     
         54 . A kit comprising the set of oligonucleotide paints of  claim 37 . 
     
     
         55 . The kit of  claim 54 , further comprising instructions for use. 
     
     
         56 . The kit of  claim 54 , wherein the kit is a diagnostic kit. 
     
     
         57 . The kit of  claim 56 , wherein the kit is used to determine karyotype of a sample. 
     
     
         58 . A kit comprising the set of oligonucleotide paints of  claim 49 . 
     
     
         59 . The kit of  claim 58 , further comprising instructions for use. 
     
     
         60 . The kit of  claim 58 , wherein the kit is a diagnostic kit. 
     
     
         61 . The kit of  claim 60 , wherein the kit is used to determine karyotype of a sample. 
     
     
         62 . A method of preparing a plurality of high resolution oligonucleotide paints comprising:
 computationally determining genomic spacing of a plurality of synthetic, oligonucleotide sequences, wherein each of the plurality is complementary to a portion of a specific chromosome sequence;   synthesizing the plurality of synthetic oligonucleotide sequences; and   labelling the plurality of synthetic oligonucleotide sequences with a detectable label to produce a plurality of oligonucleotide paints, wherein the set of oligonucleotide paints has a resolution of about two kilobases or fewer, and wherein each of a plurality of the oligonucleotide paints is complementary to a target nucleic acid sequence of 40 consecutive nucleotide bases or fewer.   
     
     
         63 . The method of  claim 62 , wherein the plurality of the oligonucleotide paints is complementary to a target nucleic acid sequence of 30 consecutive nucleotide bases or fewer. 
     
     
         64 . The method of  claim 62 , wherein the plurality of the oligonucleotide paints is complementary to a target nucleic acid sequence of 20 consecutive nucleotide bases or fewer. 
     
     
         65 . The method of  claim 62 , wherein the plurality of the oligonucleotide paints is complementary to a target nucleic acid sequence of 10 consecutive nucleotide bases or fewer. 
     
     
         66 . The method of  claim 62 , further comprising the step of computationally selecting at least one detectable label to label each of the plurality of synthetic, oligonucleotide sequences. 
     
     
         67 . The method of  claim 62 , further comprising the step of computationally determining the presence of single nucleotide polymorphisms in a genomic sequence of interest to reduce synthesis of synthetic oligonucleotide sequences that bind to repeated regions of the genomic sequence of interest. 
     
     
         68 . A method of making a set of high resolution oligonucleotide paints comprising:
 providing at least one solid support having a plurality of synthetic, single stranded oligonucleotide sequences attached thereto, wherein a portion of each of the plurality of synthetic, single stranded oligonucleotide sequences is complementary to a portion of a specific chromosome sequence;   synthesizing a plurality of complementary strands, each of which is complementary to a synthetic, single stranded oligonucleotide sequence attached to the at least one solid support;   removing the plurality of complementary strands from the at least one solid support;   amplifying the plurality of complementary strands; and   labelling the plurality of complementary strands to produce a set of high resolution oligonucleotide paints, wherein each of a plurality of the oligonucleotide paints is complementary to a target nucleic acid sequence of 40 consecutive nucleotide bases or fewer.   
     
     
         69 . The method of  claim 68 , wherein the plurality of the oligonucleotide paints is complementary to a target nucleic acid sequence of 30 consecutive nucleotide bases or fewer. 
     
     
         70 . The method of  claim 68 , wherein the plurality of the oligonucleotide paints is complementary to a target nucleic acid sequence of 20 consecutive nucleotide bases or fewer. 
     
     
         71 . The method of  claim 68 , wherein the plurality of the oligonucleotide paints is complementary to a target nucleic acid sequence of 10 consecutive nucleotide bases or fewer. 
     
     
         72 . The method of  claim 68 , wherein the plurality of oligonucleotide paints can cross a cell membrane. 
     
     
         73 . The method of  claim 68 , wherein the plurality of oligonucleotide paints can cross a nuclear membrane. 
     
     
         74 . The method of  claim 68 , further comprising the step of:
 hybridizing the oligonucleotide paints to one or more target sequences.   
     
     
         75 . The method of  claim 74 , further comprising the step of:
 extending the plurality of hybridized oligonucleotide paints.   
     
     
         76 . The method of  claim 75 , wherein the extending step is performed by primer extension. 
     
     
         77 . The method of  claim 75 , further comprising the step of washing the extended plurality of hybridized oligonucleotide paints under stringent conditions. 
     
     
         78 . The method of  claim 75 , wherein the oligonucleotide paints include a detectable label and a quencher. 
     
     
         79 . The method of  claim 78 , wherein the quencher is released during the step of extending. 
     
     
         80 . The method of  claim 78 , wherein the detectable label is a fluorescent label. 
     
     
         81 . The method of  claim 68 , further comprising the step of:
 hybridizing the oligonucleotide paints to one or more target sequences in the presence of an enzyme selected from the group consisting of one or more of a proteinase, a lipase, and a ribonuclease.   
     
     
         82 . The method of  claim 68 , wherein the target nucleic acid sequences are present in a multi-well plate. 
     
     
         83 . The method of  claim 82 , wherein the multi-well plate is a 384-well plate. 
     
     
         84 . The method of  claim 75 , wherein the hybridized oligonucleotide paints are detected by fluorescent in situ hybridization (FISH). 
     
     
         85 . The method of  claim 68 , wherein the target nucleic acid sequence is genomic.