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|Title:||Construction and Characterization of Deletion Mutations in Domain C of ARSl|
|Author(s):||Woontner, Michael Roger|
|Doctoral Committee Chair(s):||Scott, John F.,|
|Department / Program:||Microbiology|
|Degree Granting Institution:||University of Illinois at Urbana-Champaign|
|Abstract:||Autonomously Replicating Sequences, or ARS elements, promote high-frequency transformation and extrachromosomal maintenance of plasmids in Saccharomyces cerevisiae, properties expected of DNA replication origins. A series of overlapping deletions in one flanking region (Domain C) of the ARSl element was constructed, and the effect of the deletions on the maintenance of various multicopy and single-copy plasmids examined. Analysis of the stabilities and copy numbers of multicopy plasmids indicated that while the core consensus element is absolutely required for extrachromosomal maintenance, the absence of Domain C has little effect. The loss rates of centromere (single-copy) plasmids increased slightly as the deletions approached the core consensus, suggesting that a block of sequence between 225 and 255 nucleotides from the consensus contains an element important to the maximal function of ARSl. These results also suggested that ARSl plays a role in replication, but has no effect on the segregation of centromere plasmids.
Comparison of multicopy plasmids of different sizes, each with an intact ARSl element, indicated a destabilizing effect of sequences derived from the E. coli cloning vector pBR322. A small amount of pBR322 (about a kilobase) was tolerated, and the bacterial ori was not responsible for the destabilizing effect.
Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 1987.
|Date Available in IDEALS:||2014-12-16|