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  • 1
    Abstract: Here we describe a set of tools to facilitate the use of maltose and the MAL32 promoter for regulated gene expression in yeast, alone or in combination with the GAL1 promoter. Using fluorescent protein reporters we find that under non-inducing conditions the MAL32 promoter exhibits a low basal level of expression, similar to the GAL1 promoter, and that both promoters can be induced independently of each other using the respective sugars, maltose and galactose. While their repression upon glucose addition is immediate and complete, we found that the MAL32 and GAL1 promoter each exhibit distinct induction kinetics. A set of plasmids is available to facilitate the application of the MAL32 promoter for chromosomal modifications using PCR targeting and for plasmid based gene expression. This article is protected by copyright. All rights reserved.
    Type of Publication: Journal article published
    PubMed ID: 27714848
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  • 2
    Keywords: CELLS ; GROWTH ; CELL ; human ; liver ; CLONING ; GENOME ; PROTEIN ; PROTEINS ; SACCHAROMYCES-CEREVISIAE ; DNA ; FAMILY ; PROTEIN FAMILIES ; PROTEIN FAMILY ; MEMBER ; MEMBERS ; DELETION ; MUTANT ; SUBUNIT ; YEAST ; STABILITY ; PHENOTYPE ; REPLICATION ; OVEREXPRESSION ; HIGH-LEVEL ; budding yeast Saccharomyces cerevisiae,mitochondrial DNA maintenance,Mmf1p,mitochondrial large ribos ; MMF1P
    Abstract: Members of the YERO57c/YJGFc/UK114 protein family have been identified in bacteria and eukaryotes. The budding yeast Saccharomyces cerevisiae contains two different proteins of this family, Hmf1p and Mmf1p. We have previously shown that Mmf1p is a mitochondrial protein functionally related to its human homologue and able to influence the maintenance of mitochondrial DNA. Deletion of Mmf1 results in loss of the mitochondrial genome. Using a multicopy suppression approach, we have identified a protein of the mitochondrial large ribosomal subunit, MRPL40, which stabilizes mtDNA in Deltammf1 cells. Overexpression of MRPL40 did not prevent loss of mtDNA in a mutant strain lacking the mitochondrial protein Abf2p. Thus, MRPL40 does not have a general effect on mtDNA stability, but it may be specific for the mmf1-null strain. We also show that the Deltamrpl40 cells present a similar phenotype to the mmfl-null strain, having reduced mtDNA stability and growth rate. Furthermore, we observed that rho(+)Deltamrpl40 haploid cells can be obtained when tetrads are directly dissected on medium containing a non-fermentable carbon source. Thus, replication and segregation of the mtDNA can occur in the absence of MRPL40. We also show that another mitochondrial ribosomal protein, MRPL38, is able to overcome the Deltammfl-associated defect. Together, our results suggest a link between Mmf1p and the two mitochondrial ribosomal proteins. Copyright (C) 2004 John Wiley Sons, Ltd
    Type of Publication: Journal article published
    PubMed ID: 15164357
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  • 3
  • 4
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 1 (1985) 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 5
    ISSN: 0749-503X
    Keywords: Cyclic AMP ; phosphoprotein phosphatase ; protein kinase ; suppressor ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The ppd1 mutant of yeast, Saccharomyces cerevisiae, was isolated as a suppressor of the cyr2 mutation which caused alteration of the catalytic subunit of cAMP-dependent protein kinase. Three peaks of phosphoprotein phosphatase activity (peak I, II and III) were identified by DEAE-Sephacel chromatography of crude extracts of the wild-type strain. The ppd1 mutant was deficient in peak III phosphoprotein phosphatase activity. The peak III enzyme efficiently utilized the phosphorylated forms of NAD-dependent glutamate dehydrogenase and trehalase as substrate. The ppd1 mutation did not suppress the cyr1, CYR3 or ras1 ras2 mutations. The ppd1 locus was located on chromosome II and had identical characteristcs with glc1. The ppd1 mutation suppressed the G1 arrest caused by nutritional limitation, but maintained sensitivity to mating pheromone. In diploids homozygous for the ppd1 mutation, no premeiotic DNA replication and commitment to intragenic recombination occurred and no spores were formed, suggesting that the accumulation of phosphorylated proteins in the absence of one of the phosphoprotein phosphatases is required for mitosis but not for the initiation of meiosis.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
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  • 6
    ISSN: 0749-503X
    Keywords: Killer ; virus-like particles ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: L-A-E double-stranded RNA (dsRNA), when introduced into cells carrying L-A-H and M2 dsRNAs, does not eliminate the L-A-H dsRNA, but (i) L-A-E does lower the copy number of L-A-H dramatically and (ii) L-A-E eliminates M2 dsRNA from the cell. That these two effects of L-A-E are related is shown by the fact that mutants of a strain carrying L-A-H and M2 selected for their resistance to exclusion of M2 by L-A-E [effect (ii)] have an altered L-A-H whose copy number is not lowered by L-A-E [effect (i)]. Although the L-A in K1 strains (L-A-HN in all cases examined) differs significantly both genetically and physically from the L-A in the K2 strain studied (L-A-H), the L-A-HN from the K1 strains can maintain M2 dsRNA, and the L-A-H from the K2 strains can maintain M1 dsRNA.
    Additional Material: 2 Ill.
    Type of Medium: Electronic Resource
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  • 7
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 1 (1985), S. 82-82 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 8
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 2 (1986), S. 129-139 
    ISSN: 0749-503X
    Keywords: Saccharomyces cerevisiae ; regulation ; pho ; pho80 ; CEN15 ; nucleotide sequence ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The PHO80 gene, which is one of the regulatory genes exerting negative control in the pho system of Saccharomyces cerevisiae, was cloned. The 1·8 kb DNA fragment carrying the PHO80 gene was sequenced and one open reading frame large enough to encode 293 amino acids was found in the sequence. Northern blot analysis of poly(A)+-RNA isolated from cells grown under repressed and derepressed conditions revealed that (i) the size of the PHO80 message was around 1·4 kb, (ii) the expression of the PHO80 gene was not affected by the presence or absence or absence of inorganic phosphate in the medium, and (iii) the expression of the PHO80 gene was not affected by pho2, pho4, pho81, or by pho80 itself. a centromere sequence was found downstream of the PHO80 coding region.
    Additional Material: 8 Ill.
    Type of Medium: Electronic Resource
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  • 9
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 1 (1985), S. 173-175 
    ISSN: 0749-503X
    Keywords: Phosphofructokinase ; glycolysis ; alternative pathway(s) ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Additional Material: 1 Ill.
    Type of Medium: Electronic Resource
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  • 10
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 1 (1985), S. 159-171 
    ISSN: 0749-503X
    Keywords: PET18 ; temperature sensitive growth ; killer ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The basis of pleiotropy shown by the pet18 mutants of Saccharomyces cerevisiae (rho-0, KIL-0 and temperature sensitive growth) was examined by cloning the fragment which complements the defect in growth at 37°C of the pet18 mutants. the cloned DNA could complement the defect in the maintenance of the killer plasmid but did not give the cell the ability to maintain mitochondrial DNA. Sequence analysis of the cloned DNA revealed the presence of four open reading frames, at least two of which are necessary for the complementation activity. By using the cloned DNA as a probe, we found that two independent pet18 mutants have a deletion covering the entire sequence contained in the probe. From these results we predict that the traits of the pet18 mutants that concern temperature sensitivity and killer of the pet18 mutants are controlled by a separate gene(s) from that which participates in the maintenance of mitochondrial DNA.
    Additional Material: 7 Ill.
    Type of Medium: Electronic Resource
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