Präprative isolierung von desoxiriboadenylsäuren aus hydrolysaten oxidierter heringsspermen-DNA

Abstract

Preparative isolation of deoxyriboadenylic acids from hydrolysates of oxidised herring sperm DNA The preparative-scale chemical degradation of DNA from herring sperm to mixtures of purine oligonucleotides or deoxyriboadenylic acids is described. The deoxyriboadenylic acid mixture was separated into six fractions according to increasing ionic charge by column chromatography on QAE-Sephadex A-25. Of the first fraction, 75% was deoxyriboadenosine monophosphate; of the second, 99% consisted of a mixture of p(dA) 2 and (dA) 2p, while of the third 93% was pdAp. The remaining fractions contained more or less complex mixtures of longer-chain oligonucleotides which could be further separated by subsequent re-chromatography on QAE-Sephadex at pH 9.6. By application of paper chromatography to the fractions obtained from column chromatography, the pure nucleotide phosphate pdAp, p(dA) 2p, p(dA) 3p, p(dA) 4p and the mixtures of sequence isomers p(dA) 2, (dA) 2p; p(dA) 3, (dA) 3p; and p(dA) 6, (dA) 6p could be isolated preparatively. The nucleotide phosphate were converted to (dA) 2, (dA) 3 or (dA) 4 and the mixtures of sequence isomers to (dA) 2, (dA) 3 or (dA) 6 by treatment with alkaline phosphate. By this means dephosphorylated, paper chromatographically pure oligodexyriboadenylic acid may be obtained in a preparative scale from the hydrolysates. The structures of the nucleotides thus isolated were deduced from the absorbtion characteristics, the R F values and the results of enzymatic degradation.