DDAO compounds as fluorescent reference standards

• US 9,790,544 B2
• Filed: 03/10/2016
• Issued: 10/17/2017
• Est. Priority Date: 12/29/2010
• Status: Active Grant

• Current Assignee: Life Technologies Corporation (Thermo Fisher Scientific Incorporated)
• Original Assignee: Life Technologies Corporation (Thermo Fisher Scientific Incorporated)
• Inventors: Benson, Scott C., Monighetti, Cinna, Koepf, Sandy M.
• Primary Examiner(s): Riley, Jezia
• Application Number:
  US15/066,248
• Publication Number:
  US 20160258004A1
• Time in Patent Office: 586 Days
• Field of Search: 435 61, 435 911
• CPC Class Codes: C09B 15/00 Acridine dyes
• CPC Class Codes: C12Q 1/6818 involving interaction of two or more labels, e.g. resonant energy transfer
• CPC Class Codes: C12Q 1/6844 Nucleic acid amplification reactions
• CPC Class Codes: C12Q 1/6851 Quantitative amplification
• CPC Class Codes: C12Q 2527/125 Specific component of sample, medium or buffer
• CPC Class Codes: C12Q 2545/101 with an internal standard/control
• CPC Class Codes: C12Q 2561/113 Real time assay
• CPC Class Codes: C12Q 2563/107 fluorescence

Abstract

According to the present teachings, methods and compositions are provided that utilize at least one reference dye of formula (I):

In some embodiments, a method comprises measuring a detection signal of a reporter dye and at least one reference dye of formula (I). In some embodiments, a composition comprises a reference dye of formula (1), a buffer, a selection of nucleotides and a protein.

Claims

1. An amplification master mix composition comprising:

a buffer, a selection of nucleotides, at least one protein moiety, and a reference dye, wherein the reference dye has a structure according to formula (I);

2. The composition of claim 1, wherein the buffer is selected from tris, tricene, bicene, HEPES, and MOPS.

3. The composition of claim 2, wherein the buffer further comprises a chloride salt selected from potassium, lithium, magnesium and manganese, and a surfactant.

4. The composition of claim 3, wherein the surfactant is selected from a polysorbate surfactant, a polyoxyethylene surfactant, a polyethoxylated phenol surfactant, and a zwitterionic surfactant.

5. The composition of claim 1, wherein the selection of nucleotides is from deoxynucloetides (dNPTs).

6. The composition of claim 5, wherein the selection of nucleotides is further from dideoxynucleotides (didNTPs), deaza-GTP, deaza-dGTP, and 2′

-deoxyinosine 5′

-triphosphate (dITP).

7. The composition of claim 1, wherein the protein is selected from a DNA polymerase, a ligase, a reverse transcriptase, a ribonuclease, a glycosylase, a single strand binding protein, a pyrophosphatase, an albumin, and a gelatin.

8. The composition of claim 1, wherein the composition further comprises primers.

9. The composition of claim 1, wherein the composition is lyophilized.

10. A multiwell plate comprising an amplification master mix composition, wherein the composition comprises:

a buffer, a selection of nucleotides, at least one protein moiety, and a reference dye, wherein the reference dye has a structure according to formula (I);

11. The multiwell plate of claim 10, wherein the buffer selected from tris, tricene, bicene, HEPES, and MOPS.

12. The multiwell plate of claim 10, wherein the buffer further comprises a chloride salt selected from potassium, lithium, magnesium and manganese, and a surfactant.

13. The multiwell plate of claim 12, wherein the surfactant is selected from a polysorbate surfactant, a polyoxyethylene surfactant, a polyethoxylated phenol surfactant, and a zwitterionic surfactant.

14. The multiwell plate of claim 10, wherein the selection of nucleotides is from deoxynucloetides (dNPTs).

15. The multiwell plate of claim 14, wherein the selection of nucleotides is further from dideoxynucleotides (didNTPs), deaza-GTP, deaza-dGTP, and 2′

-deoxyinosine 5′

-triphosphate (dITP).

16. The multiwell plate of claim 10, wherein the protein is selected from a DNA polymerase, a ligase, a reverse transcriptase, a ribonuclease, a glycosylase, a single strand binding protein, a pyrophosphatase, an albumin, and a gelatin.

17. The multiwell plate of claim 10, wherein the composition further comprises primers.

18. The multiwell plate of claim 10, wherein composition lyophilized.