title: Covalent attachment of DNA to agarose. Improved synthesis and use in affinity chromatography.
abstract: DNA has been covalently linked to insoluble matrices of agarose (Sepharose) in high yield using cyanogen bromide activation. Both double-stranded and single-stranded DNA have been coupled with yields up to 225 nmol/mg dry weight Sepharose or 3-8 mumol nucleotide phosphate/ml bed volume. The DNA-Sepharose has been used for (a) the affinity chromatography of various enzymes (Escherichia coli DNA polymerase I and RNA polymerase) from crude extracts or after initial purification steps, resulting in high yields and degrees of purification, and for (b) nucleic acid hybridization. The DNA-Sepharose is stable to high temperature, prolonged storage, and in the case of single-stranded DNA, can be washed with NaOH to destroy nuclease activity and to release any digested oligonucleotides or mononucleotides.
mesh_terms: D000818:Animals; D001665:Binding Sites; D002417:Cattle; D002846:Chromatography, Affinity / Q000379:methods*; D003488:Cyanogen Bromide; D004247:DNA; D004254:DNA Nucleotidyltransferases / Q000302:isolation & purification; D012321:DNA-Directed RNA Polymerases / Q000302:isolation & purification; D004355:Drug Stability; D004926:Escherichia coli / Q000201:enzymology; D007700:Kinetics; D009691:Nucleic Acid Denaturation; D009693:Nucleic Acid Hybridization; D011134:Polysaccharides; D012685:Sepharose; D013696:Temperature; D013950:Thymus Gland; D013997:Time Factors
chemical_list: D011134:Polysaccharides; D004247:DNA; D012685:Sepharose; D004254:DNA Nucleotidyltransferases; D012321:DNA-Directed RNA Polymerases; D003488:Cyanogen Bromide