The isolation of duplex DNA fragments containing (dG.dC) clusters by chromatography on poly(rC)-Sephadex.
Duplex DNA containing oligo(dG.dC)-rich clusters can be isolated by specific binding to poly(rC)-Sephadex. This binding, probably mediated by the formation of an oligo(dG.dC)rC+ triple helix, is optimal at pH 5 in 50% formamide, 2 M LiCl; the bound DNA is recovered by elution at pH 7.5. Using this method we find that the viral DNAs PM2, lambda and SV40 contain at least 1, 1 and 2 sites for binding to poly(rC)-Sephadex, respectively. These binding sites have been mapped in the case of SV40; the binding sites can in turn be used for physical mapping studies of DNAs containing (dG.dC) clusters. Inspection of the sequence of the bound fragments of SV40 DNA shows that a (dG.dC)6-7 tract is required for the binding of duplex DNA to poly(rC)-Sephadex. Although about 60% of rabbit DNA cleaved with restriction endonuclease KpnI binds to poly(rC)-Sephadex, no binding is observed for the 5.1 kb DNA fragment generated by KpnI digestion, which contains the rabbit beta-globin gene. This indicates that oligo(dG.dC) clusters are not found close to the rabbit beta-globin gene.
D000818:Animals; D001483:Base Sequence; D002846:Chromatography, Affinity / Q000379:methods; D003090:Coliphages / Q000235:genetics; D004247:DNA / Q000302:isolation & purification*; D004262:DNA Restriction Enzymes; D004279:DNA, Viral / Q000032:analysis; D005796:Genes; D005914:Globins / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D009693:Nucleic Acid Hybridization; D011066:Poly C; D011817:Rabbits; D013539:Simian virus 40 / Q000235:genetics
D004279:DNA, Viral; D011066:Poly C; D005914:Globins; D004247:DNA; D004262:DNA Restriction Enzymes