Methods for nucleic acid capture

Abstract

Solutions, reagents, and methods for nucleic acid purification. In certain aspects, cationic surfactant and, optionally, an anionic surfactant solutions are provided which can be used for phase separation and capture of nucleic acids, such as plasmid or genomic DNA, to a solid phase carrier, such as a mineral matrix.

1 - 101 . (canceled) 
     
     
         102 . A method of isolating plasmid DNA, comprising:
 (a) obtaining a sample comprising plasmid DNA;   (b) capturing the plasmid DNA to a mineral matrix with a phase separation solution comprising domiphen bromide (DB) under conditions that allow for selective capture of plasmid DNA versus RNA;   (c) treating the mineral matrix and captured plasmid DNA with a salt solution, thereby enhancing the retention of the captured plasmid DNA;   (d) washing the mineral matrix and captured plasmid DNA with an organic wash solution; and   (e) eluting the plasmid DNA from the mineral matrix, thereby isolating the plasmid DNA.   
     
     
         103 . The method of  claim 102 , wherein the salt solution comprises lithium salt, sodium salt, potassium salt, magnesium salt, calcium salt, or combinations thereof. 
     
     
         104 . The method of  claim 102 , wherein the organic wash solution comprises an alcohol. 
     
     
         105 . The method of  claim 104 , wherein the organic wash solution comprises at least 60% ethanol or isopropanol. 
     
     
         106 . The method of  claim 102 , wherein the phase separation solution of (b) comprises 0.05% to 2% DB. 
     
     
         107 . The method of  claim 102 , wherein the phase separation solution of (b) comprises 0.05% to 1% DB and 0.05 M to 1.0 M lithium chloride, sodium chloride, potassium chloride, lithium acetate, sodium acetate, or potassium acetate. 
     
     
         108 . The method of  claim 102 , wherein the phase separation solution of (b) comprises a Tris-HCl buffer and 0.6 M to 1.05 M lithium chloride. 
     
     
         109 . The method of  claim 102 , wherein the obtaining a sample comprising plasmid DNA comprises obtaining a bacterial cell lysate by alkaline lysis. 
     
     
         110 . The method of  claim 109 , wherein obtaining the bacterial cell lysate comprises:
 (i) lysing cells comprising plasmid DNA with a first solution thereby generating a lysate; and   (ii) neutralizing the lysate with a second solution thereby precipitating genomic DNA and/or proteins.   
     
     
         111 . The method of  claim 110 , further comprising (iii) clearing the precipitate. 
     
     
         112 . The method of  claim 110 , wherein the first solution comprises sodium hydroxide and sodium dodecyl sulfate. 
     
     
         113 . The method of  claim 110 , wherein the second solution comprises potassium acetate and RNAse A. 
     
     
         114 . The method of  claim 113 , wherein the potassium acetate is present at a concentration of about 0.8 to about 3 M. 
     
     
         115 . The method of  claim 102 , wherein the isolated plasmid DNA is essentially free of endotoxin and/or PCR inhibitors. 
     
     
         116 . A composition comprising:
 (i) domiphen bromide (DB);   (ii) a salt selected from the group consisting of NaCl, NaoAc, KCl, KoAc, LiCl, LioAc, sodium formate, potassium formate, lithium formate, calcium chloride, and magnesium chloride; and   (iii) a mineral matrix, comprising nucleic acid molecules captured thereto.   
     
     
         117 . The composition according to  claim 116 , wherein the salt is lithium chloride (LiCl). 
     
     
         118 . The composition according to  claim 117 , further comprising a Tris-HCl buffer. 
     
     
         119 . The composition according to  claim 118 , comprising a final concentration (M) of about 0.05 M to 1.05 M LiCl. 
     
     
         120 . The composition according to  claim 116 , comprising a final concentration (w/v) of about 0.2% to about 0.5% of DB. 
     
     
         121 . The composition according to  claim 116 , wherein the mineral matrix is a silica matrix and wherein the nucleic acid molecules comprise plasmid DNA molecules.