Compositions and methods for nucleic acid extraction

Patent metadata

Field	Value
Publication number	US-9416356-B2
Application number	US-201414212857-A
Country	US
Kind code	B2
Filing date	Mar 14, 2014
Priority date	Mar 15, 2013
Publication date	Aug 16, 2016
Grant date	Aug 16, 2016

Abstract

Compositions and techniques for the extraction, enrichment and isolation of nucleic acids from fresh, fixed or fixed and embedded cells, tissues, biological materials and cellular source material using amine monomers are disclosed herein.

First claim

What is claimed is:

1. A method of isolating an extraction solution comprising nucleic acids from cellular source material, said method comprising:
a) providing i) cellular source material and ii) an aqueous extraction solution comprising an amine monomer, a chaotrope, and a detergent;
b) contacting said cellular source material with said extraction solution resulting in lysis of the cellular material and the isolation of an extraction solution comprising nucleic acids.

2. The method of claim 1 , wherein said amine monomer is a primary amine monomer.

3. The method of claim 2 , wherein said amine monomer is 2,2′-(ethylenedioxy)bis(ethylamine) (EDBE).

4. The method of claim 2 , wherein said amine monomer is 1,3-diaminopropane.

5. The method of claim 2 , wherein the concentration of amine monomer in said aqueous extraction solution is about 30% to about 50%.

6. The method of claim 1 , wherein said chaotrope is selected from one or more of the group consisting of urea, and guanidine thiocyanate (GITC).

7. The method of claim 6 , wherein the concentration of the chaotrope in said aqueous extraction solution is about 4 M to about 5 M.

8. The method of claim 7 , wherein said living cellular source material comprises a suspension of single cells.

9. The method of claim 8 , wherein said bacteria are Mycobacteria.

10. The method of claim 7 , wherein said suspension of cells comprise yeast.

11. The method of claim 1 , wherein said detergent is selected from one or more of the group consisting of, polysorbates, deoxycholate, sodium deoxycholate and sodium dodecyl sulfate (SDS), nonyl phenoxypolyethoxylethanol and polyethylene glycol.

12. The method of claim 11 , wherein the concentration of said detergent is about 8% to about 15%.

13. The method of Claim 1 , further comprising an alcohol wherein said alcohol is selected from one or more of the group consisting of ethanol and butanol.

14. The method of claim 13 , wherein the concentration of said alcohol is about 15% to about 25%.

15. The method of claim 1 , wherein said cellular source material is selected from living cellular source material and fixed cellular source material.

16. The method of claim 15 , wherein said suspension of cells comprise bacteria.

17. The method of claim 15 , wherein said fixed cellular source material comprises formalin-fixed paraffin embedded (FFPE) material.

18. The method of claim 1 , further comprising wherein said aqueous extraction solution is enzyme-free.

19. The method of claim 1 , further comprising wherein said aqueous extraction solution is protease-free.

20. The method of claim 1 , further comprising wherein said nucleic acids are purified from an extraction solution comprising nucleic acids with additional steps, the additional steps comprising i) adsorbing said nucleic acids onto silica coated magnetic particles by introducing said particles into said extraction solution containing extracted nucleic acids to create nucleic acid adsorbed silica coated magnetic particles; ii) washing said nucleic acid adsorbed silica coated magnetic particles, iii) eluting said nucleic acids from said silica coated magnetic particles into the elution solution and; iv) isolating said nucleic acids from said elution solution.

21. A composition comprising an aqueous extraction solution suitable for the extraction of nucleic acids from cellular source material, said composition comprising an amine monomer; a chaotropic reagent, a detergent and an organic solvent.

22. The composition of claim 21 , wherein said amine monomer is a primary amine monomer.

23. The composition of claim 22 , wherein said amine monomer is 2,2′-(ethylenedioxy)bis(ethylamine) (EDBE).

24. The composition of claim 22 , wherein said amine monomer is 1,3-diaminopropane.

25. The composition of claim 21 , wherein said chaotrope is selected from the group consisting of urea, and guanidine thiocyanate (GITC).

26. The composition of claim 25 , wherein the concentration of the chaotrope in said aqueous extraction solution is about 4 M to about 5 M.

27. The composition of claim 21 , wherein said detergent is selected from one or more of the group consisting of, polysorbates, deoxycholate, sodium deoxycholate and sodium dodecyl sulfate (SDS), nonyl phenoxypolyethoxylethanol and polyethylene glycol.

28. The composition of clam 21 , wherein the concentration of said detergent is about 8% to about 15%.

29. The composition of claim 21 , wherein said organic solvent is an alcohol and said alcohol is selected from the group consisting of ethanol and butanol.

30. The composition of claim 29 , wherein the concentration of said alcohol is about 15% to about 25%.

31. The composition of claim 21 , wherein the concentration of amine monomer in said aqueous extraction solution is about 30% to about 50%.

Assignees

• Abbott Molecular Inc

Inventors

• Gundling Gerard J

Classifications

• C12N15/1003 Primary

  Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor · CPC title

• C07H1/08

  from natural products · CPC title

• C07H19/16

  Purine radicals · CPC title

• C12N15/1013

  by using magnetic beads · CPC title

• C12Q1/6806

  Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay (C12Q1/6804 takes precedence) · CPC title