Isolation of nucleic acids

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.

1 - 33 . (canceled) 
     
     
         34 . A water soluble product for use in a method of extracting nucleic acid from a sample, the product comprising a plurality of positively ionisable groups, the ionisable groups being provided by a chemical species selected from the list consisting of:
 biological buffers;   polyhydroxylated amines;   histidine; and   polyhistidine.   
     
     
         35 . A product according to  claim 34  wherein the biological buffer is selected from the group consisting of:
 N-2-acetamido-2-amlnoethanesulfonic acid (ACES); 
 N-2-acetamido-2-iminodiacetic acid (ADA); 
 amino methyl propanediol (AMP); 3-1,1-dimethyl-2-hydroxyethylamino-2-hydroxy propanesulfonic acid (AMPSO); N,N-bis2-hydroxyethyl-2-aminoethanesulfonic acid (BES); 
 N,N-bis-2-hydroxyethylglycine (BICINE); 
 bis-2-hydroxyethyliminotrishydroxymethylmethane (Bis-Tris); N,3-bistrishydroxymethylmethylaminopropane (Bis-Tris Propane); 
 4-cyclohexylamino-1-butane sulfonic acid (CABS); 
 3-cyclohexylamino-1-propane sulfonic acid (CAPS); 
 3-cyclohexylamino-2-hydroxy-1-propane sulfonic acid (CAPSO); 
 2-N-cyclohexylaminoethanesulfonic acid (CHES); 
 3-N,N-bis-2-hydroxyethylamino-2 hydroxypropanesulfonic acid (DIPSO); N-2-hydroxyethylpiperazine-N-3-propanesulfonic acid (EPPS); N-2-hydroxyethylpiperazine-N-4-butanesulfonic acid (HEPBS); N-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid (HEPES); N-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-4-aminobutanesulfonic acid (TABS); N-trishydroxymethyl-methyl-3-aminopropanesulfonic acid (TAPS); 
 3-N-trishydroxymethyl-methylamino-2hydroxypropanesulfonic acid (TAPSO); N-trishydroxymethyl-methyl-2-aminoethanesulfonic acid (TES); 
 N-trishydroxymethylmethylglycine (TRICINE); 
 trishydroxymethylaminomethane (Tris); 
 polyhistidine; 
 polyhydroxylated imidazoles; 
 triethanolamine dimers and polymers; and 
 di/tri/oligo amino acids, for example Gly-Gly, Ser-Serf Gly-Gly-Gly, and Ser-Gly. 
 
     
     
         36 . A product according to  claim 34 , wherein the plurality of ionisable groups are separately attached to a polymer. 
     
     
         37 . A product according to  claim 34 , wherein the ionisable groups are polymerised, optionally by means of cross-linking reagents. 
     
     
         38 . A product for use in a method of extracting nucleic acid from a sample, wherein the product possesses a positive charge at both a first pH at which it is desired to bind nucleic acid and a second higher pH at which it is desired to release nucleic acid, the product comprising a plurality of negatively ionisable groups, the combined charge of which becomes more negative between said first pH and said second pH, such that the product is capable of binding nucleic acid at said first pH, which bound nucleic acid is released from the product at said second pH. 
     
     
         39 . A product according to  claim 38 , wherein the negatively ionisable group has a pKa between about 3 and 7, preferably between about 4 and 7. 
     
     
         40 . A product according to  claim 38 ; wherein the negatively ionisable is a carboxy group. 
     
     
         41 . A product according to  claim 38  wherein said positive charge is provided by a metal or metal oxide, preferably iron II,III oxide. 
     
     
         42 . A method for extracting nucleic acid from a sample containing nucleic acid, which method comprises:
 at a first pH, bringing the sample into contact with a material in which chemical species are immobilised on a solid phase, the chemical species having a positively ionisable nitrogen atom and at least two hydroxyl groups, wherein at least one of the hydroxyl groups is separated from the ionisable nitrogen by no more than two atoms, wherein the material has a positive charge at said first pH, such that nucleic acid is bound to the material; and   releasing the nucleic acid at a second, higher, pH at which the charge on the material is negative, neutral or less positive, wherein the positively ionisable nitrogen atom has a pKa between 4.5 and 8.5 and the release of the nucleic acid occurs under mild conditions.   
     
     
         43 . The method according to  claim 42 , wherein a plurality of the chemical species providing the ionisable groups are separately immobilised on a solid phase by covalent or ionic bonding or by adsorption. 
     
     
         44 . The method according to  claim 42 , wherein a plurality of the chemical species providing the ionisable groups are separately attached to a polymer, said polymer being immobilised on a solid phase by covalent or ionic bonding or by adsorption. 
     
     
         45 . The method according to  claim 44 , wherein the chemical species are covalently linked to a carboxylated polymer. 
     
     
         46 . The method according to  claim 42 , wherein the chemical species providing the ionisable groups are polymerised, optionally by means of cross-linking reagents. 
     
     
         47 . The method according to  claim 46 , wherein the polymer is immobilised on a solid support by covalent or ionic bonding or by adsorption. 
     
     
         48 . The method according to  claim 42 , wherein the chemical species providing the ionisable group is selected from amino methyl propanediol (AMP), N,N-bis-2-hydroxyethylglycine (BICINE), bis-2-hydroxyethyliminotrishydroxymethylmethane (Bis-Tris), 1,3-bistrishydroxymethylmethylaminopropane (Bis-Tris Propane), N-trishydroxymethylmethylglycine (TRICINE) or trishydroxymethylaminomethane (Tris). 
     
     
         49 . The method according to any one of  claims 42 , wherein the chemical species is a dialcohol amine reagent or a carbohydrate. 
     
     
         50 . The method according to  claim 49 , wherein the dialcohol amine is diethanol amine or the carbohydrate is glucosamine, polyglucosamine, chitosan or kanamycin. 
     
     
         51 . The method according to  claim 42 , wherein the chemical species is represented by the general formula [HO—(CH 2 ) n ] 2 —N—(CH 2 ) m , wherein n and m and integers between 1 and 10, and is attached to the solid phase using a silane reagent. 
     
     
         52 . The method according to  claim 51 , wherein the chemical species is 3-bis(2-hydroxyethyl)aminopropyltriethoxy silane. 
     
     
         53 . The method according to  claims 42 , wherein the chemical species providing the ionisable groups is a dimer or oligomer of Bis-Tris or comprises a plurality of Bis-Tris molecules attached to a polyacrylic acid backbone.