Cloning and manipulation of the Schizosaccharomyces pombe his7+ gene as a new selectable marker for molecular genetic studies.
We have cloned the his7+ gene of the fission yeast Schizosaccharomyces pombe by complementation of the recessive mutant allele his7-366. The his7+ gene is able to complement a mutation of the Escherichia coli hisI gene, suggesting that his7+ encodes a phosphoribosyl-AMP cyclohydrase. Subcloning experiments localize the gene to a 1.9-kb XbaI-BglII fragment. We describe the construction of plasmids to facilitate the use of his7+ as a selectable marker in S. pombe studies. Plasmid pEA2 carries his7+ cloned into the pUC18 polylinker. From either pEA2 or the original his7+ clone, pMN1, fragments carrying his7+ can be isolated using a variety of restriction enzymes for the construction of gene disruptions. Plasmid pEA500 is a cloning vector that carries his7+ and ars1, yet retains the ability to use the blue/white color screen to identify recombinants.
Apolinario
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Department of Biology, Boston College, Chestnut Hill, MA 02167.
Nocero
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Jin
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Hoffman
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CS
D003001:Cloning, Molecular; D004274:DNA, Recombinant / Q000378:metabolism; D004926:Escherichia coli; D005800:Genes, Fungal; D005816:Genetic Complementation Test; D005819:Genetic Markers; D005838:Genotype; D010957:Plasmids; D015183:Restriction Mapping; D012568:Schizosaccharomyces / Q000235:genetics*
D004274:DNA, Recombinant; D005819:Genetic Markers