Methods of localizing nucleic acids to arrays
Methods are disclosed relating to localizing nucleic acids to arrays such as silane-free arrays, and of sequencing the nucleic acids localized thereby.
1. A method of localizing a nucleic acid molecule to a solid support comprising:
providing a solid support comprising;
a surface;
a first plurality of nucleic acids immobilized on the surface, wherein the nucleic acids in the first plurality each comprises the sequence 5′-CAAGCAGAAGACGGCATACGA-3′ (SEQ ID NO: 3); and
a second plurality of nucleic acids immobilized on the surface, wherein the nucleic acids in the second plurality each comprises the sequence 5′-AATGATACGGCGACCACCGA-3′ (SEQ ID NO: 4);
providing a nucleic acid molecule comprising a sequence that hybridizes to SEQ ID NO: 3; and
hybridizing the nucleic acid molecule to at least one of the first plurality of nucleic acids immobilized on the surface, thereby localizing the nucleic acid molecule to the solid support.
2. The method of claim 1 , further comprising contacting the nucleic acid molecule hybridized to at least one of the first plurality of nucleic acids immobilized on the surface with reagents necessary to synthesize a complement of at least part of the nucleic acid molecule, wherein the complement to at least part of the nucleic acid molecule is covalently bound to the 3′ end of at least one of the nucleic acids immobilized on the surface.
3. The method of claim 2 , further comprising separating said nucleic acid molecule from said solid support.
4. The method of claim 2 , further comprising hybridizing said complement of at least part of the nucleic acid molecule to at least one of the second plurality of nucleic acids immobilized on the surface.
5. The method of claim 4 , further comprising determining the sequence of the nucleic acid molecule.
6. A method of localizing a nucleic acid molecule to a solid support comprising:
providing a solid support comprising;
a surface;
a first plurality of nucleic acids immobilized on the surface, wherein the nucleic acids in the first plurality each comprises the sequence 5′-CAAGCAGAAGACGGCATACGA-3′ (SEQ ID NO: 3); and
a second plurality of nucleic acids immobilized on the surface, wherein the nucleic acids in the second plurality each comprises the sequence 5′-AATGATACGGCGACCACCGA-3′ (SEQ ID NO: 4);
providing a nucleic acid molecule comprising a sequence that hybridizes to SEQ ID NO: 4; and
hybridizing the nucleic acid molecule to at least one of the second plurality of nucleic acids immobilized on the surface, thereby localizing the nucleic acid molecule to the solid support.
7. The method of claim 6 , further comprising contacting the nucleic acid molecule hybridized to at least one of the second plurality of nucleic acids immobilized on the surface with reagents necessary to synthesize a complement of at least part of the nucleic acid molecule, wherein the complement to at least part of the nucleic acid molecule is covalently bound to the 3′ end of at least one of the nucleic acids immobilized on the surface.
8. The method of claim 7 , further comprising separating said nucleic acid molecule from said solid support.
9. The method of claim 7 , further comprising hybridizing said complement of at least part of the nucleic acid molecule to at least one of the first plurality of nucleic acids immobilized on the surface.
10. The method of claim 9 , further comprising determining the sequence of the nucleic acid molecule.
11. The method of claim 6 , wherein the surface comprises a hydrogel.
12. The method of claim 11 , wherein the thickness of the hydrogel is less than 100 nm.
13. The method of claim 11 , wherein the hydrogel is the product of polymerizing a first comonomer with a second comonomer, wherein the first comonomer is selected from the group consisting of acrylamide, methacrylamide, hydroxyethyl methacrylate and N-vinyl pyrrolidinone, and wherein the second comonomer is a functionalized comonomer selected from the group consisting of acrylamide, acrylate, methacrylate and methacrylamide.
14. The method of claim 11 , wherein the hydrogel is the product of polymerizing acrylamide with N-(5-bromoacetamidylpentyl) acrylamide.
15. The method of claim 6 , wherein the first plurality of nucleic acids are immobilized on the surface via a linkage at their 5′ ends.
16. The method of claim 6 , wherein the second plurality of nucleic acids are immobilized on the surface via a linkage at their 5′ ends.
17. The method of claim 6 , wherein the first plurality of nucleic acids are immobilized on the surface via a linkage at their 5′ ends, and wherein the second plurality of nucleic acids are immobilized on the surface via a linkage at their 5′ ends.