Transcription fidelity and structural integrity of isolated nucleoli.
1. RNA was transcribed in vitro using isolated nucleoli, the endogenous form A RNA polymerase and mercurated UTP as one of the nucleoside triphosphate substrates. The products were isolated from endogenous nucleolar RNA sequences by chromatography on sulphydryl-Sepharose and analysed by hybridisation in vast DNA excess. The results of competition-hybridisation experiments suggested that a large proportion of the transcript was ribosomal RNA although some sequences had been transcribed from DNA of lower reiteration frequency. 2. Analyses of the constituents of nucleoli following isolation by the sonication procedure suggested that the nucleolar DNA, particularly the ribosomal cistrons, are severely degraded. Furthermore, indications were obtained that the transcription complexes were damaged and many of the nascent RNA chains appeared to have been sheared from near the growing points.
D000818:Animals; D001667:Binding, Competitive; D002466:Cell Nucleolus / Q000378:metabolism*; D004247:DNA / Q000302:isolation & purification / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D007700:Kinetics; D008970:Molecular Weight; D009693:Nucleic Acid Hybridization; D012313:RNA / Q000302:isolation & purification; D051381:Rats; D014158:Transcription, Genetic
D004277:DNA, Single-Stranded; D012313:RNA; D004247:DNA; D012321:DNA-Directed RNA Polymerases