Polyadenylation of RNA in a cell-free system from mouse myeloma cells

Abstract

Isolated nuclei from mouse myeloma cells which were active in RNA synthesis did not synthesize detectable amounts of poly(A)-containing RNA. On addition of a soluble protein extract from crude nuclei, the highly purified nuclei synthesized significant amounts of poly(A)-containing RNA, as analyzed by chromatography on poly(U)-Sepharose. The poly(A) tract was totally synthesized de novo and was indistinguishable from poly(A) synthesized in vivo. Twenty percent of the RNA polymerase II products were polyadenylated. More than 80% of the newly synthesized poly(A) was present on molecules at least partially transcribed in vitro. The transcription and polyadenylation reaction could be separated temporally and a portion (10%) of the polyadenylated RNA was released into the extra nuclear fraction. We conclude that this system carries out one RNA processing reaction, polyadenylation, faithfully.