期刊
YEAST
卷 27, 期 9, 页码 765-775出版社
WILEY
DOI: 10.1002/yea.1774
关键词
mutated loxP sites; yeast hybrids; hetherothallism; karyotypic diversity
资金
- BBSRC
- NERC
- Natural Environment Research Council [NE/B500190/1] Funding Source: researchfish
In this work, we developed molecular tools used in standard laboratory yeast strains, such as the cre-loxP system, so that they can be used with natural and industrial prototrophic yeast species. We constructed a new generation of dominant cassettes, with mutated loxP sites (loxLE and lox2272) and selectable drug markers, to create heterothallic strains and auxotrophic mutants without incurring in the risk of generating chromosomal rearrangements. We have shown that our newly developed loxLE-hphNT1-loxRE and lox2272-natNT2-lox2272 gene-disruption cassettes can be present in the yeast genome together with the widely used loxP-marker gene-loxP cassettes without any recombination between the lox sequences. Moreover, we also developed a new phleomycin-resistant Cre-expressing vector (to excise multiple markers simultaneously) and two new standard loxP deletion cassettes containing hygromicin B and cloNAT as selecatable markers. To validate these cassettes, we created heterothallic auxotrophic S. cerevisiae strains, without the risk of incurring gross chromosomal rearrangements, and we showed an example of a fitness study of intraspecific hybrids deriving from parents with different adaptations to carbon-limited resources. Copyright (C) 2010 John Wiley & Sons, Ltd.
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