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EP-4735620-A1 - METHODS AND FORMULATIONS FOR DENATURING DNA

EP4735620A1EP 4735620 A1EP4735620 A1EP 4735620A1EP-4735620-A1

Abstract

Disclosed herein are formulations and methods for denaturing DNA that utilize betaine as the denaturant. In some examples, the formulations include additives other than betaine, such as, for example, dimethyl sulfoxide (DMSO) and/or diethylene glycol (DEG).

Inventors

  • WEEKES, Dale
  • MILLER, Oliver

Assignees

  • Illumina, Inc.

Dates

Publication Date
20260506
Application Date
20240628

Claims (20)

  1. 1. A solution formulation for denaturing double-stranded DNA (dsDNA), the solution formulation comprising: betaine; and at least one organic solvent.
  2. 2. The solution formulation of claim 1, wherein the at least one organic solvent comprises diethylene glycol (DEG).
  3. 3. The solution formulation of claim 2, wherein a concentration of betaine in the solution formulation is between about IM and about 5M, and a concentration of DEG in the solution formulation is between about 3% and about 40%.
  4. 4. The solution formulation of claim 3, wherein the concentration of DEG in the solution formulation is between about 20% and about 40%.
  5. 5. The solution formulation of claim 4, wherein the concentration of DEG in the solution formulation is between about 20% and about 30%.
  6. 6. The solution formulation of any one of claims 3-5, wherein the concentration of DEG in the solution formulation is about 20%.
  7. 7. The solution formulation of any one of claims 1-6, wherein the concentration of betaine in the solution formulation is between about 2M and about 5M.
  8. 8. The solution formulation of any one of claims 1-7, wherein the concentration of betaine in the solution formulation is between about 2M and about 4M.
  9. 9. The solution formulation of any one of claims 1-8, wherein the concentration of betaine in the solution formulation is about 2M.
  10. 10. The solution formulation of any one of claims 1-8, wherein the concentration of betaine in the solution formulation is about 3M.
  11. 11. The solution formulation of claim 1, wherein the at least one organic solvent comprises diethylene glycol (DEG) and dimethyl sulfoxide (DMSO).
  12. 12. The solution formulation of claim 11, wherein a concentration of betaine in the solution formulation is between about IM and about 5M, and a concentration of the DEG in the solution formulation is between about 3% and about 40%.
  13. 13. The solution formulation of claim 11 or claim 12, wherein a concentration of DMSO in the solution formulation is between about 5% and about 30%.
  14. 14. The solution formulation of claim 13, wherein the concentration of the DMSO in the solution formulation is between about 7% and about 12%.
  15. 15. The solution formulation of claim 14, wherein the concentration of DMSO in the solution formulation is about 10%.
  16. 16. The solution formulation of claim 13, wherein the concentration of DMSO in the solution formulation is between about 17% and about 22%.
  17. 17. The solution formulation of claim 16, wherein the concentration of DMSO in the solution formulation is about 20%.
  18. 18. The solution formulation of any one of claims 11-17, wherein the concentration of the DEG in the solution formulation is between about 5% and about 25%.
  19. 19. The solution formulation of claim 18, wherein the concentration of the DEG in the solution formulation is between about 13% and about 17%.
  20. 20. The solution formulation of claim 19, wherein the concentration of the DEG in the solution formulation is about 15%.

Description

METHODS AND FORMULATIONS FOR DENATURING DNA CROSS-REFERENCE TO RELATED APPLICATIONS [0001] This application claims the benefit of U.S. Provisional Patent Application No. 63/511,076, filed June 29, 2023 and entitled “Methods and Formulations for Denaturing DNA,” the entire contents of which are incorporated by reference herein. FIELD [0002] This application relates to methods and formulations for denaturing DNA. BACKGROUND [0003] Formamide and sodium hydroxide are currently used in a number of sequencing systems to both denature the DNA library before sequencing and de-hybridise sequencing strands during clustering/resynthesis between reads However, neither formamide nor sodium hydroxide are ideal denaturing agents. Formamide is a hazardous substance. It is a suspected carcinogen and may cause birth defects. It, therefore, necessitates specialized disposal routes, hinders efforts to reach Corporate Social Responsibility goals, and is a key complaint from customers. Also, manual denaturation by NaOH increases the hands-on time of sequencing, is a potential source of human error, and requires the customer to purchase and store additional chemicals and equipment. Operator error during denaturation is a frequent issue for customers using sodium hydroxide and is a significant cause of customer run failures. Additionally, sodium hydroxide solution is hazardous to users because of its corrosive properties and cannot be used in cartridges without additional measures such as alkaline- resistant foils. SUMMARY [0004] Some examples herein provide a solution formulation for denaturing double-stranded DNA (dsDNA), the solution formulation including betaine; and at least one organic solvent. [0005] In some examples, the at least one organic solvent includes diethylene glycol (DEG). [0006] In some examples, a concentration of betaine in the solution formulation is between about IM and about 5M, and a concentration of DEG in the solution formulation is between about 3% and about 40%. [0007] In some examples, the concentration of DEG in the solution formulation is between about 20% and about 40%. In some examples, the concentration of DEG in the solution formulation is between about 20% and about 30%. In some examples, the concentration of DEG in the solution formulation is about 20%. [0008] In some examples, the concentration of betaine in the solution formulation is between about 2M and about 5M. In some examples, the concentration of betaine in the solution formulation is between about 2M and about 4M. In some examples, the concentration of betaine in the solution formulation is about 2M. In some examples, the concentration of betaine in the solution formulation is about 3M. [0009] In some examples, the at least one organic solvent includes diethylene glycol (DEG) and dimethyl sulfoxide (DMSO). [0010] In some examples, a concentration of betaine in the solution formulation is between about IM and about 5M, and a concentration of the DEG in the solution formulation is between about 3% and about 40%. [0011] In some examples, a concentration of DMSO in the solution formulation is between about 5% and about 30%. In some examples, the concentration of the DMSO in the solution formulation is between about 7% and about 12%. In some examples, the concentration of DMSO in the solution formulation is about 10%. In some examples, the concentration of DMSO in the solution formulation is between about 17% and about 22%. In some examples, the concentration of DMSO in the solution formulation is about 20%. [0012] In some examples, the concentration of the DEG in the solution formulation is between about 5% and about 25%. In some examples, the concentration of the DEG in the solution formulation is between about 13% and about 17%. In some examples, the concentration of the DEG in the solution formulation is about 15%. [0013] In some examples, the concentration of betaine in the solution formulation is between about 2M and about 3M. In some examples, the concentration of betaine in the solution formulation is about 2M. In some examples, the concentration of betaine in the solution formulation is about 2.5M. In some examples, the concentration of betaine in the solution formulation is about 3M. [0014] In some examples, the at least one organic solvent includes DMSO. [0015] In some examples, a concentration of betaine in the solution formulation is between about IM and about 5M. In some examples, the concentration of betaine in the solution formulation is between about 2M and about 3M. In some examples, the concentration of betaine in the solution formulation is about 2M. In some examples, the concentration of betaine in the solution formulation is about 2.5M. In some examples, the concentration of betaine in the solution formulation is about 3M. [0016] In some examples, a concentration of DMSO in the solution formulation is between about 5% and about 30%. In some examples, the concentration of the DMSO in the solution formulation is be