The Experts below are selected from a list of 390 Experts worldwide ranked by ideXlab platform
Wolfgang Witte - One of the best experts on this subject based on the ideXlab platform.
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aminoglycoside streptothricin resistance gene cluster aade sat4 apha 3 disseminated among multiresistant isolates of enterococcus faecium
Antimicrobial Agents and Chemotherapy, 2001Co-Authors: Guido Werner, Bianca Hildebrandt, Wolfgang WitteAbstract:Seventy-two Enterococcus faecium isolates of different origins highly resistant to Nourseothricin and streptomycin were studied. Sequencing of a genomic fragment from two isolates identified a gene cluster, aadE-sat4-aphA-3, which has been isolated recently in staphylococci and Campylobacter coli. Patterns of digested PCR products of aadE-sat4-aphA-3 were identical for all isolates.
Jürgen Wendland - One of the best experts on this subject based on the ideXlab platform.
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a molecular toolbox for manipulating eremothecium coryli
Microbiological Research, 2007Co-Authors: Selina Gastmann, Alexander Dunkler, Keith Klein, Jürgen Wendland, Andrea WaltherAbstract:Summary The genus Eremothecium contains dimorphic and filamentous fungal species, most notably Eremothecium sinecaudum (Holleya sinecauda), a dimorphic plant pathogen, which was isolated from mustard seeds, and Eremothecium gossypii (Ashbya gossypii), a filamentous fungus, which is well known for its ability to produce riboflavin. In this study, we present the initial molecular characterization of another Eremothecium species classified as Eremothecium coryli. E.coryli is a dimorphic fungus. We have developed, based on previously described reagents, a transformation system for E. coryli using kanMX and NATMX3 as dominant selectable marker genes on freely replicating plasmids conferring resistance to the antibiotics G418 and Nourseothricin, respectively. As reporter genes we could introduce lacZ and GFP, which were controlled either by the AgTEF1 promoter or by regulatable MET promoters derived from the A. gossypii and Saccharomyces cerevisiae MET3 genes. These newly established tools will allow a detailed comparison of different growth modes in filamentous or dimorphic species within the genus Eremothecium. & 2007 Elsevier GmbH. All rights reserved.
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A molecular toolbox for manipulating Eremothecium coryli.
Microbiological research, 2007Co-Authors: Selina Gastmann, Alexander Dunkler, Keith Klein, Andrea Walther, Jürgen WendlandAbstract:The genus Eremothecium contains dimorphic and filamentous fungal species, most notably Eremothecium sinecaudum (Holleya sinecauda), a dimorphic plant pathogen, which was isolated from mustard seeds, and Eremothecium gossypii (Ashbya gossypii), a filamentous fungus, which is well known for its ability to produce riboflavin. In this study, we present the initial molecular characterization of another Eremothecium species classified as Eremothecium coryli. E.coryli is a dimorphic fungus. We have developed, based on previously described reagents, a transformation system for E. coryli using kanMX and NATMX3 as dominant selectable marker genes on freely replicating plasmids conferring resistance to the antibiotics G418 and Nourseothricin, respectively. As reporter genes we could introduce lacZ and GFP, which were controlled either by the AgTEF1 promoter or by regulatable MET promoters derived from the A. gossypii and Saccharomyces cerevisiae MET3 genes. These newly established tools will allow a detailed comparison of different growth modes in filamentous or dimorphic species within the genus Eremothecium.
Guido Werner - One of the best experts on this subject based on the ideXlab platform.
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aminoglycoside streptothricin resistance gene cluster aade sat4 apha 3 disseminated among multiresistant isolates of enterococcus faecium
Antimicrobial Agents and Chemotherapy, 2001Co-Authors: Guido Werner, Bianca Hildebrandt, Wolfgang WitteAbstract:Seventy-two Enterococcus faecium isolates of different origins highly resistant to Nourseothricin and streptomycin were studied. Sequencing of a genomic fragment from two isolates identified a gene cluster, aadE-sat4-aphA-3, which has been isolated recently in staphylococci and Campylobacter coli. Patterns of digested PCR products of aadE-sat4-aphA-3 were identical for all isolates.
Ilya Borovok - One of the best experts on this subject based on the ideXlab platform.
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a gc rich prophage like genomic region of mycoplasma bovirhinis haz141_2 carries a gene cluster encoding resistance to kanamycin and neomycin
Antimicrobial Agents and Chemotherapy, 2020Co-Authors: Inna Lysnyansky, Ilya BorovokAbstract:Recently, a complete genome sequence of Mycoplasma bovirhinis HAZ141_2 was published showing the presence of a 54-kB prophage-like region. Bioinformatic analysis revealed that this region has a more than 40% GC content and a chimeric organization with three structural elements-a prophage continuous region, a restriction-modification cassette, and a highly transmittable aadE-sat4-aphA-3 gene cluster found in both Gram-positive and Gram-negative bacteria. It is known that aadE confers resistance to streptomycin, sat4 governs resistance to streptothricin/Nourseothricin, and aphA-3 is responsible for resistance to kanamycin and structurally related antibiotics. An aadE-like (aadE*) gene of strain HAZ141_2 encodes a 228-amino acid (aa) polypeptide whose carboxy-terminal domain (positions 44 to 206) is almost identical to that of a functional 302-aa AadE (positions 140 to 302). Transcription analysis of the aadE*-sat4-aphA-3 genes showed their cotranscription in M. bovirhinis HAZ141_2. Moreover, a common promoter for aadE*-sat4-aphA-3 was mapped upstream of aadE* using 5' rapid amplification of cDNA ends analysis. Determination of MICs to aminoglycosides and Nourseothricin revealed that M. bovirhinis HAZ141_2 is highly resistant to kanamycin and neomycin (≥512 μg/ml). However, MICs to streptomycin (64 μg/ml) and Nourseothricin (16 to 32 μg/ml) were similar to those identified in the prophageless M. bovirhinis type strain PG43 and Israeli field isolate 316981. We cloned the aadE*-sat4-aphA-3 genes into a low-copy-number vector and transferred them into antibiotic-sensitive Escherichia coli cells. While the obtained E. coli transformants were highly resistant to kanamycin, neomycin, and Nourseothricin (MICs, ≥256 μg/ml), there were no changes in MICs to streptomycin, suggesting a functional defect of the aadE*.
Luis G. Lugones - One of the best experts on this subject based on the ideXlab platform.
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SHORT COMMUNICATION An efficient gene deletion procedure for the mushroom-forming basidiomycete Schizophyllum commune
2016Co-Authors: Fengfeng Wang, Han A. B. Wösten, Luis G. LugonesAbstract:The Author(s) 2010. This article is published with open access at Springerlink.com Abstract Gene deletion in Schizophyllum commune is hampered by a low incidence of homologous integration. As a consequence, extensive screening is required to identify a transformant with the desired genotype. To alleviate this and to facilitate the assembly of deletion plasmids, vector pDelcas was constructed. This construct has a set of restriction sites, which allows for directional cloning of the flanking sequences at both sides of a Nourseothricin resis-tance cassette. Moreover, it contains a phleomycin resis-tance cassette elsewhere in the plasmid, which is used to screen for transformants with an ectopic integration of the pDelcas derivative. The use of pDelcas derivatives in combination with an improved PCR screening protocol permitted the efficient identification of S. commune dele-tion strains. This procedure may also function in other basidiomycetes
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an efficient gene deletion procedure for the mushroom forming basidiomycete schizophyllum commune
World Journal of Microbiology & Biotechnology, 2010Co-Authors: Robin A Ohm, Han A. B. Wösten, Jan F De Jong, Elsa Berends, Fengfeng Wang, Luis G. LugonesAbstract:Gene deletion in Schizophyllum commune is hampered by a low incidence of homologous integration. As a consequence, extensive screening is required to identify a transformant with the desired genotype. To alleviate this and to facilitate the assembly of deletion plasmids, vector pDelcas was constructed. This construct has a set of restriction sites, which allows for directional cloning of the flanking sequences at both sides of a Nourseothricin resistance cassette. Moreover, it contains a phleomycin resistance cassette elsewhere in the plasmid, which is used to screen for transformants with an ectopic integration of the pDelcas derivative. The use of pDelcas derivatives in combination with an improved PCR screening protocol permitted the efficient identification of S. commune deletion strains. This procedure may also function in other basidiomycetes.
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Phleomycin increases transformation efficiency and promotes single integrations in Schizophyllum commune.
Applied and environmental microbiology, 2008Co-Authors: Arend F. Van Peer, Charissa De Bekker, Arman Vinck, Han A. B. Wösten, Luis G. LugonesAbstract:Phleomycin is mutagenic by introducing double-strand breaks in DNA. The ble gene of Streptoalloteychus hindustanus, which confers resistance to this substance, is widely used as a selection marker for transformation. Schizophyllum commune grows on 25 μg of phleomycin ml−1 after introduction of a resistance cassette based on the ble gene. However, we here report that growth of resistant colonies on this concentration of phleomycin resulted in aberrant colony morphologies. Apparently, phleomycin was mutagenic despite acquired resistance. Therefore, a new selection system was developed based on resistance to the antibiotic Nourseothricin. However, the transformation efficiency was tenfold lower than that obtained with phleomycin as a selection agent. This low transformation efficiency could be rescued by addition of a nonselective concentration of phleomycin during protoplast regeneration. This was accompanied by a higher incidence of single-copy integrations and with an increase of expression of key genes involved in double-strand break repair. Taken together, we conclude that the effect of a nonselective concentration of phleomycin strongly resembles the effect of restriction enzyme-mediated integration (REMI) but, unlike REMI, it does not depend on the presence of a target restriction site.
