title: In vitro transcription of RNA in nuclei, nucleoli and chromatin from physarum polycephalum.
abstract: Methods for isolating nuclei, nucleoli and chromatin from Physarum polycephalum which retain high levels of endogenous RNA polymerase activity are described. Under carefully controlled conditions with respect to mono- and divalent cation concentrations RNA synthesis in nuclei displayed linear kinetics for at least 30 min and the RNA products had a similar size distribution to nuclear RNA synthesis observed in vivo. Chromatin showed 60% of the nuclear transcriptional activity but no conditions were found where faithful transcription of the template occurred. Isolated nucleoli were 5-fold more active than nuclei and the endogenous RNA polymerase activity was insensitive to alpha-amanitin. Under carefully controlled conditions, the nucleoli appeared to support the accurate transcription, re-initiation and processing of rRNA chains in vitro.
mesh_terms: D002458:Cell Fractionation; D002466:Cell Nucleolus / Q000201:enzymology; D002467:Cell Nucleus / Q000201:enzymology; D002843:Chromatin / Q000201:enzymology; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D008938:Mitosis; D009235:Myxomycetes / Q000235:genetics*; D010804:Physarum / Q000235:genetics* / Q000378:metabolism; D012313:RNA / Q000096:biosynthesis*; D014158:Transcription, Genetic
chemical_list: D002843:Chromatin; D012313:RNA; D012321:DNA-Directed RNA Polymerases