Abstract
A disruption cassette has been constructed containing the LEU2 gene flanked by directly repeated sitespecific recombination sites of the yeast plasmid, pSB3, which resembles the 2 μm DNA of Saccharomyces cerevisiae. A disruption constructed by inserting this DNA fragment acquires a Leu+ phenotype, which can be easily removed by expressing the FLP-PSB3 gene encoding the site-specific recombinase of pSB3. A test was made using a Schizosaccharomyces pombe host. The ura4 + gene of S. pombe was replaced with the ura4::LEU2 gene constructed by inserting the disruption cassette into the ura4 + gene. Then, the FLP-pSB3 gene driven by the nmt1 + promoter was introduced into this disruptant. Upon de-repression of the nmt1 promoter by removing thiamine from the medium, the rate of appearance of Leu- was increased. As expected the ura4 + locus underwent a structural change. Thus, the FLP-pSB3 protein and its target site can function adequately in S. pombe.
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Communicated by R. J. Schweyen
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Toh-e, A. Construction of a marker gene cassette which is repeatedly usable for gene disruption in yeast. Curr Genet 27, 293–297 (1995). https://doi.org/10.1007/BF00352095
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DOI: https://doi.org/10.1007/BF00352095