Isolation of poly(A)+ RNA by paper affinity chromatography

Abstract

Poly(A)+ RNA was isolated from in vitro short-term-labeled total cytoplasmic RNA of Ehrlich ascites tumor cells by oligo(dT) cellulose chromatography. This poly(A)+ RNA fraction was compared with a poly(A)+ RNA fraction isolated by a new procedure which involves specific binding of poly(A)+ RNA to messenger affinity paper (mAP) and its release in hot (70 degrees C) water. In typical experiments 10-11 micrograms (2.3%) of poly(A)+ RNA can be retained from 500 micrograms of total cytoplasmic RNA per cm2 of mAP in a quick one-step procedure. The poly(A)+ RNA preparations isolated by the two methods proved to be almost identical with respect to their fraction in total cytoplasmic RNA, specific radioactivities, sucrose gradient profiles, and translation assays. Since the isolation of poly(A)+ RNA by mAP is much less time consuming than that by oligo(dT) column chromatography and since the poly(A)+ RNA can be recovered from mAP in small volumes, which avoids further loss during precipitations, it can be advantageously used for preparative isolation of poly(A)+ RNA.