Compositions and methods for preparing oligonucleotide solutions
The present invention is directed to methods and compositions for generating a pool of oligonucleotides. The invention finds use in preparing a population or subpopulations of oligonucleotides in solution. The pool of oligonucleotides finds use in a variety of nucleic acid detection and/or amplification assays.
1. A method of detecting the presence of target nucleic acids comprising:
a) obtaining a first substrate comprising greater than 50 different single-stranded oligonucleotides linked to discrete sites on the first substrate;
b) releasing said greater than 50 different single stranded oligonucleotides from said first substrate as a pool of greater than 50 different single stranded oligonucleotides;
c) contacting said pool of greater than 50 different single stranded oligonucleotides with a composition comprising target nucleic acids, whereby said target nucleic acids hybridize with greater than 50 different single stranded oligonucleotides in said pool of oligonucleotides;
d) capturing said greater than 50 different single stranded oligonucleotides in said pool of oligonucleotides hybridized with said target nucleic acids on a second substrate;
e) contacting said target nucleic acids with a primer, wherein said target nucleic acids are amplified upon binding to said primer; and
f) detecting the presence of said target nucleic acids.
2. The method of claim 1 , wherein step b) further comprises amplifying said greater than 50 different single stranded oligonucleotides that are released from said first substrate, wherein said releasing and amplifying generates said pool of greater than 50 different single stranded oligonucleotides.
3. The method of claim 2 , wherein said amplifying in step b) comprises polymerase chain reaction.
4. The method of claim 2 , wherein said amplifying in step b) comprises rolling circle amplification.
5. The method of claim 1 , wherein said discrete sites are contiguous with other sites.
6. The method of claim 1 , wherein said discrete sites are non-contiguous with other sites.
7. The method of claim 1 , wherein said greater than 50 different single stranded oligonucleotides that are linked to said discrete sites on the first substrate are each linked through a cleavable linker.
8. The method of claim 7 , wherein said releasing of said greater than 50 single stranded oligonucleotides from said first substrate comprises cleavage of each said cleavable linker with light.
9. The method of claim 7 , wherein said releasing of said greater than 50 single stranded oligonucleotides from said first substrate comprises cleavage of each said cleavable linker with a chemical.
10. The method of claim 7 , wherein said releasing of said greater than 50 single stranded oligonucleotides from said first substrate comprises cleavage of each said cleavable linker with an enzyme.
11. The method of claim 1 , wherein said greater than 50 different oligonucleotides in said pool of oligonucleotides are labeled.
12. The method of claim 1 , wherein said first substrate comprises glass or silica.
13. The method of claim 1 , wherein said first substrate comprises plastic.
14. The method of claim 1 , wherein said first substrate is selected from the group consisting of polysaccharides, nylon, nitrocellulose, resins, silicon, carbon, and metals.