Methods and devices for nucleic acid sequence determination
View Patent ↗Methods of the invention comprise methods and devices for nucleic acid sequence determination. Generally, the invention relates to preparing a substrate for sequencing a target nucleic acid.
1. A method for conducting a chemical reaction on a surface, the method comprising the steps of:
attaching a plurality of first nucleic acid molecules via covalent linkages to reactive chemical groups on a surface;
blocking the surface using a blocking solution comprising potassium phosphate;
conducting a chemical reaction between said first nucleic acid molecules and second reactant molecules comprising an optically detectable label wherein said conducting step comprises exposing said nucleic acid to a primer under conditions sufficient to extend said nucleic acid by at least one base;
rinsing unincorporated optically labeled nucleotides from the surface; and
observing individually optically resolvable labels incorporated in the one or more first nucleic acids on the surface.
2. The method of claim 1 , wherein said first nucleic acids are DNA or RNA.
3. The method of claim 1 , wherein said optically detectable label is a fluorescent molecule.
4. The method of claim 3 , wherein said fluorescent molecule is selected from the group consisting of Cyanine-3 and Cyanine-5.
5. The method of claim 1 , wherein the reactive chemical groups are epoxide groups.
6. The method of claim 1 , wherein the first nucleic acid molecules are attached to the reactive chemical groups indirectly by means of a biotin-streptavidin binding pair wherein biotin is covalently attached to the surface.
7. The method of claim 1 , further comprising the step of exposing the surface to a drying agent.
8. The method of claim 7 , wherein said drying agent is PRB, EtOH, air or N 2 .
9. The method of claim 1 , further comprising determining nucleic acid sequences of one or more of the first nucleic acid molecules.
10. The method of claim 9 , wherein said determining step comprises conducting sequencing by synthesis.
11. The method of claim 1 , wherein the surface comprises a silica coating containing exposed epoxide groups.
12. The method as in claim 1 , wherein pH of the blocking solution is 8.5.
13. A method for conducting a chemical reaction on a surface, the method comprising the steps of:
attaching a plurality of first nucleic acid molecules via covalent linkages to reactive chemical groups on an analytical surface;
blocking the surface using a blocking solution comprising potassium phosphate;
conducting a chemical reaction using second reactant molecules comprising an optically detectable label thereby incorporating said second reactant molecules;
rinsing unincorporated optically labeled second reactant molecules from the surface; and
observing individually optically resolvable labels on the surface.