Open AccessCovalent Attachment of DNA to Agarose
Author(s) -
ARNDTJOVIN Donna J.,
JOVIN Thomas M.,
BÄHR Wolfgang,
MARQUARDT Maria,
FRISCHAUF AnneMarie
Publication year - 1975
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1975.tb04151.x
Subject(s) - sepharose , cyanogen bromide , agarose , oligonucleotide , chemistry , nucleic acid , nuclease , dna , affinity chromatography , covalent bond , dna polymerase , chromatography , dna polymerase i , biochemistry , polymerase , rna , enzyme , reverse transcriptase , organic chemistry , gene , peptide sequence
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 μmol 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.