ArticleMolecular biotechnology2011
A new method for repeated "self-cloning" promoter replacement in Saccharomyces cerevisiae.
Article in Molecular biotechnology, 2011. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 2 papers.
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The abstract states no effect estimate the extractor could read, or names no intervention and outcome on the map, so this paper lights no cell and moves no belief. It is still indexed, cited and linked below.
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Who cites it
2 citing papers in PubMed, 7 citations in OpenAlex.
- Insertion orientation within the cassette affects gene-targeting success during ends-out recombination in the yeast Saccharomyces cerevisiae.Current genetics · 2022Article
- Enhanced acetate ester production of Chinese liquor yeast by overexpressing ATF1 through precise and seamless insertion of PGK1 promoter.Journal of industrial microbiology & biotechnology · 2014Article
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Authors and funding
5 authors at 3 institutions in 3 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
A method for repeated PCR-mediated promoter replacement in the yeast Saccharomyces cerevisiae is described. It was proposed to use the DNA fragment comprising the marker gene that enables both positive and negative selection (a selectable/counter-selectable marker) surrounded by direct repeats of the desired promoter as a promoter replacement cassette. This fragment is integrated upstream of the target gene because of PCR-added terminal sequences for homologous recombination with the target locus. Subsequent marker excision via homologous recombination between the copies of the two promoters leaves one copy of the desired promoter upstream of the target genes, without any heterologous scar sequence. To test this method, a set of plasmids bearing the S. cerevisiae URA3 gene surrounded by two copies of the ADH1 or PGK1 promoter was constructed. Using these cassettes, the native promoters of the GSH1 and GSH2 genes were replaced in the ura3Δ0 recipient strains. The proposed method is useful for research applications due to simple marker excision, and for construction of "self-cloning" industrial strains, because no heterologous DNA is retained in the genome of the resulting strain after marker excision.
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