Base compositional analysis of nanogram quantities of unlabeled nucleic acids.

After conversion of unlabeled DNA and RNA to 3′-mononucleotides accurate base compositional analysis can be performed on as little as 10 ng of the hydrolysate. The 3′-mononucleotides are first quantitatively postlabeled with γ-³²P]ATP by T4 polynucleotide kinase and are then separated as mononucleoside diphosphates on Whatman DE-81 ion-exchange paper at pH 3.5 after hydrolysis of surplus γ-³²P]ATP to ³²P₁. The locations of the four labeled nucleoside diphosphates are determined by autoradiography and the ratio of radioactivity in the four spots gives the base ratio of the sample.