Ribosomal RNA genes (rDNA) exist in yeast both as a single chromosomal array of tandemly repeated units and as extrachromosomal units named 3um plasmids, although the relationship between these two forms is unclear. Inheritance of rDNA was studied using two systems. The first used a naturally occuring rDNA restriction enzyme polymorphism between two strains to distinguish between their rDNA arrays, and the second involved cloning a tRNA suppressor gene into rDNA to label individual rDNA units. An added interest to the study of the inheritance of rDNA in yeast was the possible association between it and the inheritance of the Psi factor, an enigmatic type of nonsense suppressor in yeast which shows extra-chromosomal inheritance. In a cross heterozygous for the rDNA polymorphism and the psi factor, tetrad analysis suggested that the psi factor had segregated 4:0. The majority of the rDNA units segregated in a 2:2 fashion, which suggested that reciprocal recombination in the rDNA of psi<sup>+</sup> diploids is heavily suppressed as was previously shown for psi<sup>-</sup> diploids. A heterologous plasmid containing the tRNA suppressor gene was constructed and transformed into haploid and diploid hosts. A series of transformants was obtained and physical and genetic analysis suggested that they contained tRNA suppressor gene(s) integrated into their rDNA. In a cross heterozygous for rDNA-tRNA gene insert(s), 6% of the tetrads dissected showed a meiotic segregation of the suppressed phenotype which could most probably be accounted for by inter-chromosomal gene conversion. This observation could be interpreted in two ways. Firstly, recombination intermediates between rDNA on homologues may occur in meiosis, but they are mostly resolved as gene conversions without reciprocal cross-over. Alternatively, gene conversion tracts in rDNA are rare but very long so that the tRNA gene insert was always included in the event. 3um rDNA plasmids containing the tRNA gene marker were not detected in any of the transformants analysed. An extensive quantitative analysis of the rate of reversion of the suppressed phenotype amongst these transformants identified a particulary unstable transformant group. It was proposed that the mechanism of reversion was loss of the tRNA gene insert by unequal sisterstrand exchange, and the mechanism was shown to be independent of the recombination/repair genes RAD1, RAD52, and RAD51. A genetic analysis of stability suggested that there may have been at least two loci segregating in the host strains with additive effects on stability.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:259921 |
| Date | January 1994 |
| Creators | Faridani, Jacqueline |
| Contributors | Chan, Hong-Mo |
| Publisher | University of Oxford |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://ora.ox.ac.uk/objects/uuid:71be8f51-cab2-48dc-bb7c-fb0274851c97 |
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