Methods and compositions for performing analytical operations

• US 8,956,816 B2
• Filed: 06/04/2013
• Issued: 02/17/2015
• Est. Priority Date: 06/05/2012
• Status: Active Grant

• Current Assignee: Pacific Bioscience of California Incorporated (L'Oreal SA)
• Original Assignee: Pacific Bioscience of California Incorporated (L'Oreal SA)
• Inventors: Fedorov, Andrei, Yue, Stephen, Sun, Lei, Shen, Gene, Lyle, John
• Primary Examiner(s): Noakes, Suzanne M
• Assistant Examiner(s): LACOURCIERE, GERARD MICHAEL
• Application Number:
  US13/909,797
• Publication Number:
  US 20140017674A1
• Time in Patent Office: 623 Days
• Field of Search: None
• US Class Current: 435/6.1
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: G01N 33/54393 Improving reaction conditions or stability, e.g. by coating or irradiation of surface, by reduction of non-specific binding, by promotion of specific binding

Abstract

Methods for performing analytical reactions and compositions for use in such methods, where the methods have reduced signal levels deriving from non-specific adsorption of detected reagents to other components of the analytical method, e.g., other reagents, solid phase components, vessels, etc.

Claims

1. A method of conducting an analytical reaction, said method comprising:

providing a reaction mixture comprising a first reaction component coupled to a surface of a solid support and a second reaction component having a detectable property, wherein the first reaction component comprises a polymerase; and

conducting said analytical reaction in the presence of;

(a) a first additive that reduces interpulse duration (IPD) wherein said first additive comprises a member selected from cholesterol, a cholic acid derivative, poly dT oligonucleotides, cyclodextrin, and a cyclodextrin derivative,

(b) a second additive that reduces non-specific interactions between the first reaction component and the surface, wherein the second additive comprises a member selected from an organic solvent, dimethylformamide (DMF), and N-formylmorpholine (FMP),

(c) a third additive that reduces polymerase pausing, wherein the third additive comprises a member selected from a compound comprising an aromatic ring and two —

NR₂ substituents, and an amino acid.

2. The method of claim 1, wherein said cholic acid derivative is taurocholic acid.

3. The method of claim 1, wherein said amino acid is arginine.

4. The method of claim 1, wherein said second additive is DMF.

5. The method of claim 1, wherein said cyclodextrin derivative is a sulfated cyclodextrin having the structure:

6. The method of claim 5, wherein R₁ is a sulfate group.

7. The method of claim 5, wherein R₂ and R₃ are selected from alkyl, acetamide, hydroxyalkyl, acyl, aryl or heteroaryl groups.

8. The method of claim 7, wherein R₂ and R₃ are selected from CH₂-acyl, diethylacetamide, dipropylacetamide, morpholino, piperazine, piperidine, pyrrolidine, and oxazolidine.

9. The method of claim 1, wherein the surface of the solid support further comprises a plurality of first reaction components, and wherein the plurality of first reaction components are provided disposed on the surface of the solid support as single molecules or single molecular complexes, each of which is resolvable from others of the first reaction components disposed on the solid support.

10. The method of claim 1, wherein the solid support comprises a silica based substrate.

11. The method of claim 1, wherein the second reaction component comprises a fluorescent labeling moiety.

12. The method of claim 1, wherein the second reaction component comprises a nucleotide analog.

13. The method of claim 1, wherein the organic solvent is dimethyl sulfoxide (DMSO) or dimethylacetamide (DMA).

14. The method of claim 1, wherein the compound comprising an aromatic ring and two —

NR₂ substituents is N,N-dimethyl-m-phenylenediamine (DMMP) or N,N-dimethyl-p-phenylenediamine (DMPP).

15. The method of claim 1, wherein said second additive is FMP.