Isolation of nucleic acids
First Claim
1. A product comprising a solid phase for use in a method in which the solid phase reversibly binds nucleic acid present in a sample, the product comprising a plurality of positively ionizable groups, wherein the ionizable groups are immobilized on the solid phase and are effective at a first pH at which the ionizable groups are positively charged to bind nucleic acid present in a sample and are effective to release the nucleic acid at a second, higher, pH at which the charge on the ionizable groups is negative, neutral or less positive, the ionizable groups being provided by a chemical species selected from the group consisting of biological buffers, polyhydroxylated amines, histidine and polyhistidine, wherein said plurality of positively ionizable groups have a pKa between about 4.5 and about 8.5.
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Accused Products
Abstract
A method for extracting nucleic acids from a biological material such as blood comprises contacting the mixture with a material at a pH such that the material is positively charged and will bind negatively charged nucleic acids and then eluting the nucleic acids at a pH when the said materials possess a neutral or negative charge to release the nucleic acids. The nucleic acids can be removed under mildly alkaline conditions to the maintain integrity of the nucleic acids and to allow retrieval of the nucleic acids in reagents that are immediately compatible with either storage or analytical testing.
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Citations
42 Claims
- 1. A product comprising a solid phase for use in a method in which the solid phase reversibly binds nucleic acid present in a sample, the product comprising a plurality of positively ionizable groups, wherein the ionizable groups are immobilized on the solid phase and are effective at a first pH at which the ionizable groups are positively charged to bind nucleic acid present in a sample and are effective to release the nucleic acid at a second, higher, pH at which the charge on the ionizable groups is negative, neutral or less positive, the ionizable groups being provided by a chemical species selected from the group consisting of biological buffers, polyhydroxylated amines, histidine and polyhistidine, wherein said plurality of positively ionizable groups have a pKa between about 4.5 and about 8.5.
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10. A water soluble product for use in a method of extracting nucleic acid from a sample, the product comprising a plurality of positively ionizable groups, the ionizable groups being provided by a biological buffers;
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wherein the plurality of ionizable groups are (i) separately attached to a polymer or (ii) polymerized, optionally by means of cross-linking reagents or (iii) separately attached to a polymer and polymerized, optimally by means of cross-linking reagents; and
wherein said plurality of positively ionizable groups have a pKa between about 4.5 and 8.5 and the biological buffer is selected from the group consisting of;N-2-acetamido-2-aminoethanesulfonic acid (ACES);
N-2-acetamido-2-iminodiacetic acid (ADA);
N,N-bis2-hydroxyethyl-2-aminoethanesulfonic acid (BES);
N,N-bis-2-hydroxyethylglycine (BICINE);
bis-2-hydroxyethyliminotrishydroxymethylmethane (Bis-Tris);
1,3-bistrishydroxymethylmethylaminopropane (Bis-Tris Propane);
3-N,N-bis-2-hydroxyethylamino-2-hydroxypropanesulfonic acid (DIPSO);
-2-hydroxyethylpiperazine-N-3-propanesulfonic acid (EPPS);
-2-hydroxyethylpiperazine-N-4-butanesulfonic acid (HEPBS);
-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid (HEPES);
-2-hydroxyethylpiperazine-N-2-propanesulfonic acid (HEPPSO);
2-N-morpholinoethanesulfonic acid (MES);
4-N-morpholinobutanesulfonic acid (MOBS);
3-N-morpholinopropanesulfonic acid (MOPS);
3-N-morpholino-2-hydroxypropanesulfonic acid (MOPSO);
piperazine-N-N-bis-2-ethanesulfonic acid (PIPES);
piperazine-N-N-bis-2-hydroxypropanesulfonic acid (POPSO);
N-trishydroxymethyl-methyl-3-aminopropanesulfonic acid (TAPS);
3-N-trishydroxymethyl-methylamino-2-hydroxypropanesulfonic acid (TAPSO);
N-trishydroxymethyl-methyl-2-aminoethanesulfonic acid (TES);
N-trishydroxymethylmethylglycine (TRICINE);
trishydroxymethylaminomethane (Tris);
polyhydroxylated imidazoles; and
triethanolamine dimers and polymers. - View Dependent Claims (39)
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11. A product comprising a solid phase for use in which the solid phase reversibly binds nucleic acid present in a sample, the product comprising a plurality of positively ionizable groups, wherein the ionizable groups are immobilised on the solid phase and are capable at a first pH at which the ionizable groups are positively charged of binding nucleic acid present in a sample and are capable of releasing the nucleic acid at a second, higher, pH at which the charge on the ionizable groups is negative, neutral or less positive, wherein said plurality of positively ionizable groups have a pKa between about 4.5 and about 8.5, the ionizable groups being provided by a biological buffer selected from the group consisting of:
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N-2-acetamido-2-aminoethanesulfonic acid (ACES);
N-2-acetamido-2-iminodiacetic acid (ADA);
N,N-bis2-hydroxyethyl-2-aminoethanesulfonic acid (BES);
N,N-bis-2-hydroxyethylglycine (BICINE);
bis-2-hydroxyethyliminotrishydroxymethylmethane (Bis-Tris);
1,3-bistrishydroxymethylmethylaminopropane (Bis-Tris Propane);
3-N,N-bis-2-hydroxyethylamino-2-hydroxypropanesulfonic acid (DIPSO);
-2-hydroxyethylpiperazine-N-3-propanesulfonic acid (EPPS);
-2-hydroxyethylpiperazine-N-4-butanesulfonic acid (HEPBS);
-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid (HEPES);
-2-hydroxyethylpiperazine-N-2-propanesulfonic acid (HEPPSO);
2-N-morpholinoethanesulfonic acid (MES);
4-N-morpholinobutanesulfonic acid (MOBS);
3-N-morpholinopropanesulfonic acid (MOPS);
3-N-morpholino-2-hydroxypropanesulfonic acid (MOPSO);
piperazine-N-N-bis-2-ethanesulfonic acid (PIPES);
piperazine-N-N-bis-2-hydroxypropanesulfonic acid (POPSO);
N-trishydroxymethyl-methyl-3-aminopropanesulfonic acid (TAPS);
3-N-trishydroxymethyl-methylamino-2-hydroxypropanesulfonic acid (TAPSO);
N-trishydroxymethyl-methyl-2-aminoethanesulfonic acid (TES);
N-trishydroxymethylmethylglycine (TRICINE);
trishydroxymethylaminomethane (Tris);
polyhydroxylated imidazoles; and
triethanolamine dimers and polymers. - View Dependent Claims (12, 13, 14, 15, 16, 17, 18, 19, 40)
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- 20. A product comprising a solid phase for use in which the solid phase reversibly binds nucleic acid present in a sample, the product comprising a plurality of positively ionizable groups, wherein the ionizable groups are immobilised on the solid phase and are capable at a first pH at which the ionizable groups are positively charged of binding nucleic acid present in a sample and are capable of releasing the nucleic acid at a second, higher, pH at which the charge on the ionizable groups is negative, neutral or less positive, the ionizable groups being provided by bis-2-hydroxyethyliminotrishydroxymethylmethane (Bis-Tris) and having a pKa between about 4.5 and 8.5.
- 28. A product comprising a solid phase for use in which the solid phase reversibly binds nucleic acid present in a sample, the product comprising a plurality of positively ionizable groups, wherein the ionizable groups are immobilised on the solid phase and are capable at a first pH at which the ionizable groups are positively charged of binding nucleic acid present in a sample and are capable of releasing the nucleic acid at a second, higher, pH at which the charge on the ionizable groups is negative, neutral or less positive, the ionizable groups being provided by bis-2-hydroxyethyliminotrishydroxymethylmethane (Bis-Tris) and having a pKa between about 4.5 and 8.5. and the solid phase comprises beads.
Specification