Budd, Martin E. and Campbell, Judith L. (1993) DNA polymerases delta and epsilon are required for chromosomal replication in Saccharomyces cerevisiae. Molecular and Cellular Biology, 13 (1). pp. 496-505. ISSN 0270-7306. http://resolver.caltech.edu/CaltechAUTHORS:BUDmcb93
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Three DNA polymerases, alpha, delta, and epsilon are required for viability in Saccharomyces cerevisiae. We have investigated whether DNA polymerases epsilon and delta are required for DNA replication. Two temperature-sensitive mutations in the POL2 gene, encoding DNA polymerase epsilon, have been identified by using the plasmid shuffle technique. Alkaline sucrose gradient analysis of DNA synthesis products in the mutant strains shows that no chromosomal-size DNA is formed after shift of an asynchronous culture to the nonpermissive temperature. The only DNA synthesis observed is a reduced quantity of short DNA fragments. The DNA profiles of replication intermediates from these mutants are similar to those observed with DNA synthesized in mutants deficient in DNA polymerase alpha under the same conditions. The finding that DNA replication stops upon shift to the nonpermissive temperature in both DNA polymerase alpha- and DNA polymerase epsilon- deficient strains shows that both DNA polymerases are involved in elongation. By contrast, previous studies on pol3 mutants, deficient in DNA polymerase delta, suggested that there was considerable residual DNA synthesis at the nonpermissive temperature. We have reinvestigated the nature of DNA synthesis in pol3 mutants. We find that pol3 strains are defective in the synthesis of chromosomal-size DNA at the restrictive temperature after release from a hydroxyurea block. These results demonstrate that yeast DNA polymerase delta is also required at the replication fork.
|Additional Information:||Copyright © 1993 by the American Society for Microbiology. Received 29 July 1992/Returned for modification 15 September 1992/Accepted 19 October 1992 This study was supported by grant GM 25508. We are greatly indebted to Akio Sugino for the POL2 gene disruption used in the screening for temperature-sensitive mutants.|
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|Deposited On:||03 Apr 2006|
|Last Modified:||26 Dec 2012 08:49|
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