Celniker, S. E. and Sweder, K. and Srienc, F. and Bailey, J. E. and Campbell, J. L. (1984) Deletion mutations affecting autonomously replicating sequence ARS1 of Saccharomyces cerevisiae. Molecular and Cellular Biology, 4 (11). pp. 2455-2466. ISSN 0270-7306. http://resolver.caltech.edu/CaltechAUTHORS:CELmcb84
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DNAs that contain specific yeast chromosomal sequences called ARSs transform Saccharomyces cerevisiae at high frequency and can replicate extrachromosomally as plasmids when introduced into S. cerevisiae by transformation. To determine the boundaries of the minimal sequences required for autonomous replication in S. cerevisiae, we have carried out in vitro mutagenesis of the first chromosomal ARS described, ARS1. Rather than identifying a distinct and continuous segment that mediates the ARS+ phenotype, we find three different functional domains within ARS1. We define domain A as the 11-base-pair (bp) sequence that is also found at most other ARS regions. It is necessary but not sufficient for high-frequency transformation. Domain B, which cannot mediate high-frequency transformation, or replicate by itself, is required for efficient, stable replication of plasmids containing domain A. Domain B, as we define it, is continuous with domain A in ARS1, but insertions of 4 bp between the two do not affect replication. The extent of domain B has an upper limit of 109 bp and a lower limit of 46 bp in size. There is no obvious sequence homology between domain B of ARS1 and any other ARS sequence. Finally, domain C is defined on the basis of our deletions as at least 200 bp flanking domain A on the opposite side from domain B and is also required for the stability of domain A in S. cerevisiae. The effect of deletions of domain C can be observed only in the absence of domain B, at least by the assays used in the current study, and the significance of this finding is discussed.
|Additional Information:||Copyright © 1984 by the American Society for Microbiology. Received 4 June 1984/Accepted 7 August 1984 This investigation was supported by Public Health Service grant GM 25508 from the National Institutes of Health and by grants from The American Cancer Society (MV-142), The March of Dimes (1-794), The National Science Foundation (CPE-8118425), and Energy Conservation and Utilization Technology, Program of the U.S. Department of Energy. J.L.C. is the recipient of RCDA CA 00544.|
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