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Biomedical subjects

D Ramón

Publications and source records attributed to D Ramón.

At least 55 records · Page 3Linked to original sources
PubMed1994

Development of a transformation system for Trichoderma longibrachiatum and its use for constructing multicopy transformants for the egl1 gene.

An efficient transformation system for the fungus Trichoderma longibrachiatum has been developed. Transformation was obtained both by electroporation and polyethyleneglycol treatment, using a plasmid carrying the Escherichia coli hygromycin B phosphotransferase gene as a dominant selectable marker. The transformation frequency was 0.5 to 5 transformants/micrograms plasmid DNA. Transformation normally occurred by tandem integration of the transforming DNA. A high percentage of the transformants were mitotically unstable. The efficiency of co-transformation was very high (around 90%), and several co-transformants containing multiple copies of the egl1 gene encoding a beta-(1,4)-endoglucanase were obtained. Some of them secrete increased levels of endoglucanase to the culture medium. In addition, the E. coli lacZ gene was expressed in an active form under control of the Aspergillus nidulans gpdA gene promoter.

Base Sequence↗
PubMed1994

Population dynamics of natural Saccharomyces strains during wine fermentation.

Using mitochondrial DNA restriction endonuclease analysis, the dynamics of the natural Saccharomyces cerevisiae strains present in spontaneous wine fermentations have been studied. We observed a sequential substitution of Sacch. cerevisiae strains along fermentation agreeing with different fermentation phases. When the restriction pattern's similarity (measured as the fraction of shared restriction fragments) was high, a clear sequential substitution of the strains was seen. However, when the similarity was low, although a sequential substitution could be also observed between secondary strains, a clearly predominant strain was present along the whole fermentation process.

Colony Count, Microbial↗
PubMed1994

The Bacillus subtilis lipoprotein LplA causes cell lysis when expressed in Escherichia coli.

A gene called lplA (lipoprotein-like) has been isolated from a genomic library of Bacillus subtilis expressed in Escherichia coli. Clones carrying the lplA gene were selected by the ability of the colonies to give visible haloes of starch hydrolysis. The cloned fragment contains an open reading frame (ORF) of 1509 bp encoding a protein of 56 kDa. The protein contains a typical N-terminal signal sequence, a putative transmembrane anchor domain and a leucine zipper at the C-terminus. The expression of this protein in E. coli causes cell lysis, only the N-terminal domain of the LplA protein being responsible for this phenotype. The mechanism of cell lysis is similar to that previously suggested for the expression in E. coli of the lipoproteins encoded by the Streptococcus pneumoniae genes malX and amiA. The protein is modified with palmitic acid when secreted in E. coli, confirming that it is a typical lipoprotein.

Amino Acid Sequence↗
PubMed1993

Arabinan degrading enzymes from Aspergillus nidulans: induction and purification.

The presence in Aspergillus nidulans of two enzymes related to the Aspergillus niger endo-arabinase and alpha-L-arabinofuranosidase B has been established using antibodies against the purified A. niger enzymes. Moreover, the absence of an equivalent in A. nidulans to the alpha-L-arabinofuranosidase A of A. niger has been confirmed both at the protein and at the DNA level. Both A. nidulans arabinases have been purified and physico-chemically and kinetically characterized. They have a much higher temperature optimum than the corresponding A. niger enzymes. The pattern of induction has been studied on media containing different carbon sources showing an important role of L-arabitol in the induction of these enzymes.

Aspergillus nidulans↗
PubMed1993

Nucleotide sequence of a Trichophyton mentagrophytes HindIII mitochondrial DNA fragment containing a tRNA gene cluster.

A 0.85-kb HindIII mitochondrial DNA fragment of the dermatophytic fungus Trichophyton mentagrophytes has been sequenced. The fragment contains eight complete genes which corresponds to a tRNA gene cluster. From 5' to 3', the sequenced genes code for tRNA(thr), tRNA(glu), tRNA(val), tRNA(met1), tRNA(met3), tRNA(leu), tRNA(ala), and tRNA(phe). This tRNA gene cluster is located downstream of the larger ribosomal RNA gene. The particularities of the sequenced genes and their comparison with other fungal tRNA mitochondrial genes are reported.

Base Sequence↗
PubMed1993

Construction of a recombinant wine yeast strain expressing beta-(1,4)-endoglucanase and its use in microvinification processes.

A genetic transformation system for an industrial wine yeast strain is presented here. The system is based on the acquisition of cycloheximide resistance and is a direct adaptation of a previously published procedure for brewing yeasts (L. Del Pozo, D. Abarca, M. G. Claros, and A. Jiménez, Curr. Genet. 19:353-358, 1991). Transformants arose at an optimal frequency of 0.5 transformant per microgram of DNA, are stable in the absence of selective pressure, and produce wine in the same way as the untransformed industrial strain. By using this transformation protocol, a filamentous fungal beta-(1,4)-endoglucanase gene has been expressed in an industrial wine yeast under the control of the yeast actin gene promoter. Endoglucanolytic wine yeast secretes the fungal enzyme to the must, producing a wine with an increased fruity aroma.

Base Sequence↗
PubMed1993

[Utilization of molecular techniques for the characterization of wine yeasts and the study of the wine-making process].

The study of the fermentation process in the Alicante region allowed us to conclude that adverse climatic conditions could be responsible for deficient wine-making with serious problems arising fermentations, usually causing incomplete fermentation. In order to avoid these problems, we selected a Saccharomyces cerevisiae strain, namely T73, isolated in the same Alicante region, to be used to perform controlled fermentations. The use of selected strains in the wine-making process requires the development of characterization techniques that can clearly differentiate between the inoculated strain and the wild strains present in the musts. In order to differentiate strains present in the wine ecosystems, an extensive survey of different methods of yeast strains identification has been carried out. However, these techniques are very complex to be used in industry. For this reason, we have developed a new, simple, inexpensive and rapid method based on mitochondrial DNA restriction analysis. This technique was applied to the control of wine fermentations conducted by active dry yeasts. This molecular approach allows us to understand the role of the inoculated dry yeast strain and that of the natural S. cerevisiae flora during wine fermentation.

DNA, Fungal↗
PubMed1992

Isolation of an Aspergillus terreus mutant impaired in arginine biosynthesis and its complementation with the argB gene from Aspergillus nidulans.

Using filtration enrichment techniques, an Aspergillus terreus arginine auxotrophic strain which contains a mutation that abolishes ornithine transcarbamylase (OTCase) activity has been isolated. This mutant has been genetically transformed with the cloned Aspergillus nidulans OTCase gene. Prototrophic transformants arose at a frequency of about 50 transformants per microgram of plasmid DNA. Southern blot analysis of DNA from the transformants showed that the transforming DNA was ectopically integrated at different locations in the A. terreus genome, often in multiple tandem copies. The transformants were phenotypically stable for several mitotic divisions and retained their capacity to produce extracellular enzymes.

Arginine↗
PubMed1992

Cloning, sequence analysis and yeast expression of the egl1 gene from Trichoderma longibrachiatum.

A gene (egl1) encoding an endoglucanase (EGL1) from Trichoderma longibrachiatum has been cloned and sequenced. This gene, homologous to the T. reesei egl1 gene, differs from it in the length of the introns (particularly the first one) and encoded protein. A cDNA fragment obtained by the rapid amplification of cDNA ends method, which takes advantage of the polymerase chain reaction, has been expressed in yeast under control of the cyc-gal inducible promoter and yeast clones able to secrete active enzyme have been obtained.

Amino Acid Sequence↗
PubMed1992

Molecular monitoring of wine fermentations conducted by active dry yeast strains.

A simple and rapid method of yeast strain characterization based on mitochondrial DNA restriction analysis was applied to the control of wine fermentations conducted by active dry yeast strains. This molecular approach allows us to understand several important aspects of this process, such as the role of the active dry yeast strain and that of the natural Saccharomyces cerevisiae flora during vinification. In this paper, we demonstrate that the inoculated strain is really responsible for the fermentation but does not suppress significant development of natural strains during the first stages. During this early period, natural strains could have important effects on wine flavor.

Journal Article↗
PubMed1991

Transformation of Aspergillus terreus with the hygromycin B resistance marker from Escherichia coli.

Aspergillus terreus was transformed to hygromycin B resistance using a bacterial resistance gene under the control of Aspergillus nidulans regulatory sequences. Southern hybridization of transformants indicated that in most of the cases the vector DNA was integrated into the recipient chromosome in the form of tandem arrays. Transformants were mitotically stable in both selective and non-selective medium and retained their capacity to produce xylanase or glucoamylase activities.

Aspergillus↗
PubMed1991

Xanthelasmoid mastocytosis.

An 8-year-old girl had small, papular vulval lesions for six years; the lesions were yellowish with numerous surface depressions. Symptoms due to the action of mastocyte mediators were observed. A biopsy specimen showed a dense monomorphous infiltrate of the deep dermis by rounded cells with granular cytoplasm and a round or oval central nucleus. The morphology of the lesions and red-purple metachromatic staining led to the diagnosis of xanthelasmoid mastocytosis. Symptoms were controlled with hydroxyzine. Annual follow-up has shown no evidence of systemic involvement to date. Surgery should be contemplated as a future therapeutic option, in view of the location of the lesions.

Child↗
PubMed1990

Sequences and homology analysis of two genes encoding beta-glucosidases from Bacillus polymyxa.

The nucleotide sequences of the bglA and bglB genes encoding beta-glucosidases from Bacillus polymyxa have been determined. Both genes contain coding regions of 1344 bp, corresponding to polypeptides with Mrs of 51,643 and 51,547, respectively. Patterns of codon usage indicate that both genes are expressed at a low frequency. Previous data suggested that the proteins encoded by bglA and bglB were intra- and extracellular enzymes, respectively; however, neither of the two deduced amino acid sequences has N termini with the typical features of a leader peptide. The proteins encoded by bglA and bglB show remarkable homology to each other and to other beta-glucosidases (Bgl) and beta-galactosidases (beta Gal). On the basis of the observed homologies, we can define two groups of microbial Bgl: one of them, type I, including most bacterial Bgl, and type II, including enzymes from different yeast species and one from Clostridium thermocellum. Likewise, at least two groups of beta Gal can be distinguished: type I, including enzymes homologous to type-I Bgl, and type II, showing no homology to any of the previous groups.

Amino Acid Sequence↗
PubMed1990

Sequences of isopenicillin N synthetase genes suggest horizontal gene transfer from prokaryotes to eukaryotes.

Evolutionary distances between bacterial and fungal isopenicillin N synthetase (IPNS) genes have been compared to distances between the corresponding 5S rRNA genes. The presence of sequences homologous to the IPNS gene has been examined in DNAs from representative prokaryotic organisms and Ascomycotina. The results of both analyses strongly support two different events of horizontal transfer of the IPNS gene from bacteria to filamentous fungi. This is the first example of such a type of transfer from prokaryotes to eukaryotes.

Acremonium↗