An efficient transformation procedure enabling long‐term storage of competent cells of various yeast genera

Dohmen, R. Jürgen; Strasser, Alexander W.M.; Honer, Christian; Hollenberg, Cornelis P.

doi:10.1002/yea.320070704

Cited by 367 publications

(243 citation statements)

References 8 publications

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“…For plates 2% Bacto-agar was added. Standard methods were used for genetic analysis (Campbell and Duffus 1988), yeast transformation (Dohmen et al 1991) and Escherichia coil transformation (Sambrook et al 1989). …”

Section: Methodsmentioning

confidence: 99%

Sequence of the HAP3 transcription factor of Kluyveromyces lactis predicts the presence of a novel 4-cysteine zinc-finger motif

Mulder¹,

Scholten²,

Boer³

et al. 1994

Molec. Gen. Genet.

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Disclaimer/Complaints regulationsIf you believe that digital publication of certain material infringes any of your rights or (privacy) interests, please let the Library know, stating your reasons. In case of a legitimate complaint, the Library will make the material inaccessible and/or remove it from the website. Please Ask the Library: http://uba.uva.nl/en/contact, or a letter to: Library of the University of Amsterdam, Secretariat, Singel 425, 1012 WP Amsterdam, The Netherlands. You will be contacted as soon as possible. Abstract The Kluyveromyces lactis homologue of the Saecharomyces cerevisiae HAP3 gene was isolated by functional complementation of the respiratorydeficient phenotype of the S. cerevisiae hap3::HIS4 strain SHY40. The K1HAP3 gene encodes a protein of 205 amino acids, of which the central B-domain of 90 residues is highly homologous to HAP3 counterparts of S. cerevisiae and higher eukaryotes. The protein contains a novel 4-cysteine zinc-finger motif and we propose by analogy that all other homologous HAP3 proteins contain the same motif, with the position containing the third cysteine being occupied by a serine residue. In contrast to the situation in S. cerevisiae, disruption of the K1HAP3 gene in K. lactis does not result in a respiratory-deficient phenotype and the growth of the null strain is indistinguishable from wild type. There is also no effect on the expression of the carbon source-regulated KICYC1 gene, suggesting either a different role for the HAP2/3/4 complex, or the existence of a different mechanism of carbon source regulation. Sequence verification of the S. cerevisiae HAP3 locus reveals that, just as in K. lactis, a long open reading frame (ORF) is present upstream of the HAP3 gene. These highly homologous ORFs are predicted to have at least eight membrane-spanning fragments, but do not show significant homology to any known sequence present in databases. The ScORFX gene is transcribed in the opposite direction to ScHAP3, but, in contrast to an earlier report by Hahn et al. (1988), the transcripts of the two genes do not overlap. The model proposed by these authors, in which the ScHAP3 gene is regulated by an anti-sense non-coding mRNA, is therefore not correct.

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Section: Methodsmentioning

confidence: 99%

Sequence of the HAP3 transcription factor of Kluyveromyces lactis predicts the presence of a novel 4-cysteine zinc-finger motif

Mulder¹,

Scholten²,

Boer³

et al. 1994

Molec. Gen. Genet.

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“…(1989). Yeast transformation was performed by the method of Dohmen et al (1991). Yeast cells were grown at 30°C either in liquid culture with vigorous shaking or on agar plates (media supplemented with 18 g agar I-1).…”

Section: Strains Growth Conditions and Plasmidsmentioning

confidence: 99%

Characterization of a novel eukaryotic ATP/ADP translocator located in the plastid envelope of Arabidopsis thaliana L.

et al. 1997

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SummaryRecently, we have sequenced a cDNA clone from Arabidopsis thaliana L. encoding a novel putative ATP/ADP translocator (AATP1). Here, we demonstrate that the radioactively labeled AATP1 precursor protein, synthesized in vitro, is targeted to envelope membranes of isolated spinach chloroplasts. Antibodies raised against a synthetic peptide of AATP1 recognized a single polypeptide of about 62 kDa in chloroplast inner envelope preparations. The cDNA coding for the AATP1 protein was functionally expressed in Saccharomyces cerevisiae and Escherichia coil In both expression systems, increased rates of ATP transport were observed after reconstitution of the extracted protein into proteoliposomes. To our knowledge, this is the first report on the functional expression of an intrinsic plant membrane protein in E. coil To yield high rates of ATP transport, proteoliposomes had to be preloaded with ADP, indicating a counter-exchange mode of transport. Carboxyatractyloside did not substantially interfere with ATP transport into proteoliposomes containing the plastidic ATP/ADP translocator. An apparent KM for ATP of 28 llM was determined which is similar to values reported for isolated plastidso The data presented here strongly support the conclusion that AATP1 represents a novel eukaryotic adenylate carrier and that it is identical with the so far unknown plastidic ATP/ADP translocator.

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“…uvarum species. We then tested the method described by Dohmen et al (1991) that uses frozen competent cells. In our hands, this method was shown to be efficient, and we thus used it in all further experiments.…”

Section: Genetic Tools and Methodsmentioning

confidence: 99%

“…S. bayanus var. uvarum cell transformation was performed using frozen competent cells (Dohmen et al, 1991). Genomic DNA from S. bayanus var.…”

Section: Dna Manipulationsmentioning

confidence: 99%

Developing methods and strains for genetic studies in the Saccharomyces bayanus var. uvarum species

Talarek¹,

Louis²,

Cullin³

et al. 2004

Yeast

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For years, Saccharomyces cerevisiae has been used as a model organism to gain insight into complex biological processes. The study of closely related yeast species may be critical for understanding the molecular mechanism of evolution. Among those species, S. bayanus var. uvarum could be particularly pertinent because of the availability of its genome sequence. However, to date, in that species genetic studies are problematical due to the lack of standard strains collection and genetic methods. Here, we have developed heterothallic S. bayanus var. uvarum strains and obtained stable haploid strains. We further used UV-induced mutation and gene disruption to create a collection of auxotrophic derivatives. Finally, we have elaborated or improved methods to cultivate cells, obtain zygotes and spores and to transform this species. All these tools can now be used by the scientific community to study the biology of this species.

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An efficient transformation procedure enabling long‐term storage of competent cells of various yeast genera

Cited by 367 publications

References 8 publications

Sequence of the HAP3 transcription factor of Kluyveromyces lactis predicts the presence of a novel 4-cysteine zinc-finger motif

Sequence of the HAP3 transcription factor of Kluyveromyces lactis predicts the presence of a novel 4-cysteine zinc-finger motif

Characterization of a novel eukaryotic ATP/ADP translocator located in the plastid envelope of Arabidopsis thaliana L.

Developing methods and strains for genetic studies in the Saccharomyces bayanus var. uvarum species

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