A system for the analysis of yeast ribosomal DNA mutations.

AUTOR(ES)

Musters, W

RESUMO

To develop a system for the analysis of eucaryotic ribosomal DNA (rDNA) mutations, we cloned a complete, transcriptionally active rDNA unit from the yeast Saccharomyces cerevisiae on a centromere-containing yeast plasmid. To distinguish the plasmid-derived ribosomal transcripts from those encoded by the rDNA locus, we inserted a tag of 18 base pairs within the first expansion segment of domain I of the 26S rRNA gene. We demonstrate that this insertion behaves as a neutral mutation since tagged 26S rRNA is normally processed and assembled into functional ribosomal subunits. This system allows us to study the effect of subsequent mutations within the tagged rDNA unit on the biosynthesis and function of the rRNA. As a first application, we wanted to ascertain whether the assembly of a 60S subunit is dependent on the presence in cis of an intact 17S rRNA gene. We found that a deletion of two-thirds of the 17S rRNA gene has no effect on the accumulation of active 60S subunits derived from the same operon. On the other hand, deletions within the second domain of the 26S rRNA gene completely abolished the accumulation of mature 26S rRNA.

Documentos Relacionados

• Isolation and analysis of multicopy extragenic suppressors of dnaA mutations.
• Induction of yeast killer factor mutations.
• Mutational analysis of the lambda int gene: DNA sequence of dominant mutations.
• Lambda YES: a multifunctional cDNA expression vector for the isolation of genes by complementation of yeast and Escherichia coli mutations.
• Modification of enzymatically amplified DNA for the detection of point mutations.