Parameters of Xenopus rDNA Transcription in Microinjected Oocytes
Microinjection experiments using cloned Xenopus laevis rDNA have resulted in a detailed analysis of transcriptional control elements of the rDNA repeat (Busby and Reeder, 1983; Labhart and Reeder, 1984; De Winter and Moss, 1986). However, to facilitate detection of X. laevis rDNA transcripts most microinjection experiments were performed in a heterologous system: X. laevis rDNA clones were injected into X. borealis oocytes. It is clear that this assay has to cope with the phenomenon of nucleolar dominance of X. laevis transcripts over X. borealis transcripts (Reeder and Roan, 1984). Another line of evidence from oocyte microinjection experiments indicated, that oocyte batches from different females are likely to contain different amounts of rDNA specific transcription factors (Trendelenburg et al., 1978; Sollner-Webb and McKnight, 1982; McStay and Reeder, 1986). From EM-observations of injected rDNA templates it is clear that transcription factors specific for RNA polymerase I compete with those for polII on individual injected templates (Trendelenburg and Gurdon, 1978; Trendelenburg and Puvion-Dutilleul, 1987). In order to allow quantitation of some essential parameters influencing rDNA transcription we used a homologous experimental system: we injected a X. laevis rDNA construct of which transcripts could be discerned from endogenous rDNA transcripts into X. laevis oocytes and monitored the amount of transcript produced by the S1 nuclease technique.
KeywordsEDTA Formamide Blin Amanitin
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