Formulations for nucleic acid stabilization on solid substrates

Patent metadata

Field	Value
Publication number	US-9040675-B2
Application number	US-201213721948-A
Country	US
Kind code	B2
Filing date	Dec 20, 2012
Priority date	Apr 30, 2012
Publication date	May 26, 2015
Grant date	May 26, 2015

Abstract

The present disclosure generally relates to dry solid matrices for the extraction, stabilization, and storage of nucleic acids, particularly RNA, in a dry format under ambient conditions for a prolonged period of time. Methods for collecting and recovering the nucleic acids stored in the dry solid matrix are also described.

First claim

The invention claimed is:

1. A dry solid matrix for extraction and storage of nucleic acids from a biological sample, wherein a composition comprising;
a metal thiocyanate comprises a Group 1 or Group 2 metal cation;
a reducing agent where the reducing agent is tris(2-carboxyethyl)phosphine (TCEP); and
a buffer; is incorporated into the dry solid matrix, and the matrix is then dried; and where the solid matrix is a porous matrix comprising cellulose, cellulose acetate, glass fiber, or any combination thereof.

2. The dry solid matrix of claim 1 , wherein the Group 1 or Group 2 metal cation is selected from the group consisting of Na + , K + , Li + , Mg +2 , Ca +2 , and Ba +2 .

3. The dry solid matrix of claim 1 , wherein the composition present in the solid matrix further comprises a UV inhibitor, a free-radical trap, a chelator, or any combination thereof

4. The dry solid matrix of claim 1 , wherein the composition incorporated into the dry solid matrix further comprises an RNase inhibitor.

5. The dry solid matrix of claim 1 , wherein the dry solid matrix permits prolonged storage of nucleic acids in a dry format under ambient conditions.

6. The dry solid matrix of claim 5 , wherein the nucleic acids are RNA, DNA, or a combination thereof.

7. The dry solid matrix of claim 6 , wherein the nucleic acids are RNA.

8. The dry solid matrix of claim 1 , wherein the porous matrix is a cellulose paper.

9. The dry solid matrix of claim 1 , wherein the metal thiocyanate is selected from the group consisting of sodium thiocyanate, potassium thiocyanate, lithium thiocyanate, magnesium thiocyanate, calcium thiocyanate, barium thiocyanate, zinc thiocyanate, and a combination thereof.

10. The dry solid matrix of claim 1 , wherein the buffer is selected from the group consisting of 2-amino-2-hydroxymethyl-propane-1,3-diol (Tris), 2-(N-morpholino)ethanesulfonic acid (MES), 3-(N-morpholino)propanesulfonic acid (MOPS), 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES), a citrate buffer, and a phosphate buffer.

11. The dry solid matrix of claim 10 , wherein the pH range is between 3 and 6.

12. The dry solid matrix of claim 3 , wherein the UV protectant or free-radical trap is selected from the group consisting of hydroquinone monomethyl ether (MEHQ), hydroquinone (HQ), toluhydroquinone (THQ), and ascorbic acid.

13. The dry solid matrix of claim 4 , wherein the RNase inhibitor is vanadyl ribonucleoside complex (VRC) or a commercially available RNase inhibitor.

14. The solid matrix of claim 13 further comprising a free-radical trap, wherein the free-radical trap comprises MEHQ or THQ.

15. A method for extracting and storing nucleic acids from a sample comprising:
a) providing a dry solid matrix for extraction and storage of nucleic acids from a biological sample, where the solid matrix is a porous matrix comprising cellulose, cellulose acetate, glass fiber, or any combination thereof, and wherein a composition is incorporated into the dry solid matrix comprising,
a metal thiocyanate comprises a Group 1 or Group 2 metal cation;
a reducing agent ,where the reducing agent is TCEP;
and a buffer;
b) drying the solid matrix after incorporation of the composition into the dry solid matrix;
c) applying a sample to the dry solid matrix to collect the nucleic acids;
d) drying the solid matrix comprising the nucleic acids; and
e) storing the nucleic acids on the solid matrix in a dry state under ambient conditions.

16. The method of claim 15 , wherein the metal cation is selected from the group consisting of Na + , K + , Li + , Mg +2 , Ca +2 , and Ba +2 .

17. The method of claim 15 , wherein the method further comprises recovering the nucleic acids from the solid matrix.

18. The method of claim 15 , wherein the biological sample is blood, serum, tissue, saliva, nasal mucous, or cells.

19. The method of claim 15 , wherein the sample is a purified nucleic acid sample or a tissue culture cell preparation.

20. The method of claim 15 , wherein purification of the nucleic acids from the sample is not required prior to applying the sample to the dry solid matrix for extraction and storage of the nucleic acids.

21. The method of claim 15 , wherein the method permits the prolonged storage of RNA in a dry format under ambient conditions.

22. The method of claim 17 , wherein the nucleic acids are recoved from the dry solid matrix by rehydrating the matrix in an aqueous solution, a buffer, or an organic solution, and wherein the nucleic acids are subjected further analysis.

23. The method of claim 17 , wherein the nucleic acids are recovered from the dry solid matrix by electroelution.

Assignees

• Gen Electric

Inventors

• Bales Brian Christopher
• Kvam Erik Leeming
• Davis Jason Louis

Classifications

• G01N1/40

  Concentrating samples · CPC title

• C12N15/1006

  by means of a solid support carrier, e.g. particles, polymers · CPC title

• B01J20/045 Primary

  containing sulfur, e.g. sulfates, thiosulfates, gypsum · CPC title

• B01J20/24

  Naturally occurring macromolecular compounds, e.g. humic acids or their derivatives · CPC title

• C12N15/1017

  by filtration, e.g. using filters, frits, membranes · CPC title