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  • Transformation  (6)
  • Cyanobacteria  (5)
  • Springer  (11)
  • Public Library of Science (PLoS)
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  • Springer  (11)
  • Public Library of Science (PLoS)
Years
  • 1
    Electronic Resource
    Electronic Resource
    Transformation of Aspergillus flavus: construction of urate oxidase-deficient mutants by gene disruption (1992)
    Springer
    Current genetics 21 (1992), S. 447-453 
    Details
    ISSN: 1432-0983
    Keywords: Aspergillus flavus ; Urate oxidase ; Transformation ; Gene disruption
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A transformation procedure based on the complementation of a genetic defect was developed using a nitrate reductase-deficient mutant of Aspergillus flavus. The initial transformation efficiency was improved 40-fold by combining factors in a planned experimental program. Although low, this transformation rate was sufficient to obtain transformants in which the urate oxidase-encoding gene (uaZ) was disrupted in a gene replacement experiment. These new uaZ- strains were unable to utilize uric acid as the unique nitrogen source and could be reversed directly to the wild-type phenotype in second order transformation experiments using a urate oxidase-expressing vector.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00351654
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  • 2
    Electronic Resource
    Electronic Resource
    Characterization of competent cells and early events of Agrobacterium-mediated genetic transformation in Arabidopsis thaliana (1992)
    Springer
    Planta 188 (1992), S. 439-456 
    Details
    ISSN: 1432-2048
    Keywords: Agrobacterium ; Arabidopsis ; Cotyledon ; Root explant (in vitro culture) ; Transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The insertion of foreign DNA in plants occurs through a complex interaction between Agrobacteria and host plant cells. The marker gene β-glucuronidase of Escherichia coli and cytological methods were used to characterize competent cells for Agrobacterium-mediated transformation, to study early cellular events of transformation, and to identify the potential host-cell barriers that limit transformation in Arabidopsis thaliana L. Heynh. In cotyledon and leaf explants, competent cells were mesophyll cells that were dedifferentiating, a process induced by wounding and-or phytohormones. The cells were located either at the cut surface or within the explant after phytohormone pretreatment. In root explants, competent cells were present in dedifferentiating pericycle, and were produced only after phytohormone pretreatment. Irrespective of their origin, the competent cells were small, isodiametric with thin primary cell walls, small and multiple vacuoles, prominent nuclei and dense cytoplasm. In both cotyledon and root explants, histological enumeration and β-glucuronidase assays showed that the number of putatively competent cells was increased by preculture treatment, indicating that cell activation and cell division following wounding were insufficient for transformation without phytohormone treatment. Exposure of explants for 48 h to A. tumefaciens produced no characteristic stress response nor any gradual loss of viability nor cell death. However, in the competent cell, association between the polysaccharide of the host cell wall and that of the bacterial filament was frequently observed, indicating that transformation required polysaccharide-to-polysaccharide contact. Flow cytofluorometry and histological analysis showed that abundant transformation required not only cell activation (an early state exhibiting an increase in nuclear protein) but also cell proliferation (which in cotyledon tissue occurred at many ploidy levels). Noncompetent cells could be made competent with the appropriate phytohormone treatments before bacterial infection: this should aid analysis of critical steps in transformation procedures and should facilitate developing new strategies to transform recalcitrant plants.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00192812
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  • 3
    Electronic Resource
    Electronic Resource
    Role of the host cell cycle in theAgrobacterium-mediated genetic transformation ofPetunia: Evidence of an S-phase control mechanism for T-DNA transfer (1997)
    Springer
    Planta 201 (1997), S. 160-172 
    Details
    ISSN: 1432-2048
    Keywords: Agrobacterium ; Leaf mesophyll cells ; Petunia ; Transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Chimeric β-glucuronidase (GUS) gene expression in an efficientAgrobacterium-mediated transformation system utilising mesophyll cells ofPetunia hybrida synchronized with cell cycle phase-specific inhibitors (mimosine and colchicine) was used to show the absolute requirement of S-phase for transfer and/or integration of the transferred DNA (T-DNA). Flow-cytometric analysis of nuclear DNA content and immunohistological detection of bromodeoxyuridine (BrdUrd) incorporation showed that, prior to phytohormone treatment, most (98%) mesophyll cells were at GO-Gl-phase (quiescent phase) and no cell division was occurring. After 48 h and 72 h of phytohormone treatment, there was a rapid increase in S-G2-M-phase populations (〉 75%) and a concomitant decrease (down to 24%) in G0–-G1-phase cells. Assays of GUS showed that maximum transformation (〉 95% of explants) also occurred after this period. Our data showed that mimosine and colchicine blocked the mesophyll cells at late Gl-phase and M-phase, respectively. No transformation (= GUS expression) was observed in phytohormone-treated cells inhibited in late G1 by mimosine. However, after removal of mimosine, 82% of the explants were transformed, indicating the non-toxic and reversible effect of the inhibitor. On the other hand, a relatively high transformation frequency (65% of explants) was observed after blocking the cell cycle at M-phase with colchicine. However, only transient, but no stable, gene expression (= kanamycin-resistant callus formation) was observed in colchicine-treated M-phase-arrested cells. Similarly, endoreduplication of nuclear DNA, which occurred during the 48 h of phytohormone treatment in some mesophyll cells and cells located along the minor veins in the leaf explants, resulted in transient GUS expression only. These observations indicate a direct correlation between endoreduplication and transient GUS gene expression. Obviously, for stable GUS gene expression, cell division and proliferation are required, indicating that both DNA duplication (S-phase) and cell division (M-phase) are strongly related to stable transformation. We propose that the present system should facilitate further dissection of the process of T-DNA integration in the host genome and therefore should aid in developing new strategies for transformation of recalcitrant plants.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF01007700
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  • 4
    Electronic Resource
    Electronic Resource
    High efficiency transformation of Tolypocladium geodes conidiospores to phleomycin resistance (1991)
    Springer
    Current genetics 20 (1991), S. 309-314 
    Details
    ISSN: 1432-0983
    Keywords: Phleomycin resistance ; Transformation ; Fungi
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A convenient and efficient transformation system has been developed for the filamentous fungus Tolypocladium geodes. In contrast to most of the commonly described techniques requiring prior preparation of protoplasts or spheroplasts, this method leads to high efficiency transformation of T. geodes conidiospores following moderate lytic enzyme treatment. Competent cells so obtained are still resistant to osmotic pressure and can be stored frozen without loss of viability. The highest transformation frequency (3-5x103 transformants per μg of DNA) was obtained with plasmid pUT737 containing the Sh ble gene conferring phleomycin resistance under the control of a strong promoter isolated from Trichoderma reesei. Southern hybridization revealed multiple integration sites of plasmid DNA into the T. geodes nuclear DNA despite the absence of homology between the transforming DNA and the recipient genome. Instability could not be detected for the phleomycin® phenotype during more than five generations of mitotic growth under non-selective conditions.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00318520
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  • 5
    Electronic Resource
    Electronic Resource
    A method to sharply delimit a yeast nuclear gene in a cloned DNA fragment (1982)
    Springer
    Current genetics 6 (1982), S. 159-162 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Transformation ; Gene subcloning
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We have developped a procedure to delimit the boundaries of a cloned gene carried on a DNA fragment as large as 4 to 5 kilobases. The method consists in the following. Two series of limit digest products generated with a tetranucleotide recognition sequence endonuclease and originating from either of the two ends of this DNA segment are tested for their complementing capacity by yeast transformation. The gene is then delimited by the overlap of the two shortest complementing fragments.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00435216
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  • 6
    Electronic Resource
    Electronic Resource
    Structure and mutation of a gene encoding a Mr 33000 phycocyanin-associated linker polypeptide (1990)
    Springer
    Archives of microbiology 153 (1990), S. 541-549 
    Details
    ISSN: 1432-072X
    Keywords: Cyanobacteria ; Synechococcus sp. PCC 7002 ; Agmenellum quadruplicatum ; Phycobilisome ; Phycocyanin ; Linker polypeptide ; Photosynthetic antenna ; Nucleotide sequence ; Directed mutagenesis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The gene encoding a phycocyanin-associated linker polypeptide of Mr 33000 from the cyanobacterium Synechococcus sp. PCC 7002 was found to be located adjacent and 3′ to the genes encoding the α and β subunits of phycocyanin. The identity of this gene, designated cpcC, was proven by matching the amino-terminal sequence of the authentic polypeptide with that predicted by the nucleotide sequence. A cpcC mutant strain of this cyanobacterium was constructed. The effect of the mutation was to prevent assembly of half the total phycocyanin into phycobilisomes. By electron microscopy, phycobilisomes from this mutant were shown to contain rod substructures composed of a single disc of hexameric phycocyanin, as opposed to two discs in the wild type. It was concluded that the Mr 33000 linker polypeptide is required for attachment of the core-distal phycocyanin hexamer to the core-proximal one. Using absorption spectra of the wild type, CpcC−, and phycocyanin-less phycobilisomes, the in situ absorbances expected for specific phycocyanin-linker complexes were calculated. These data confirm earlier findings on isolated complexes regarding the influence of linkers on the spectroscopic properties of phycocyanin.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00245263
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  • 7
    Electronic Resource
    Electronic Resource
    The structure of Gloeobacter violaceus and its phycobilisomes (1981)
    Springer
    Archives of microbiology 129 (1981), S. 181-189 
    Details
    ISSN: 1432-072X
    Keywords: Cyanobacteria ; Gloeobacter violaceus ; Synechocystis ; Freeze-etching ; Membranes ; Phycobilisomes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The fine structure of the atypical cyanobacterium Gloeobacter violaceus has been studied on frozen-etched replicas and compared to that of a typical unicellular strain: Synechocystis 6701. The complementary fracture faces of G. violaceus cytoplasmic membrane contain particles less numerous and more heterogenous in size than either the cytoplasmic membrane or the thylakoid membranes of Synechocystis. The most frequently observed particles of the exoplasmic fracture (EF) face of the G. violaceus cytoplasmic membrane are 11 nm in diameter and occasionally form short alignments. This particle class is similar in appearance to the numerous, aligned EF particles of Synechocystis thylakoid membranes. In replicas of cross-fractured G. violaceus, a layer 50–70 nm thick, composed of rod-like elements, underlies the inner surface of the cytoplasmic membrane. The rods, 12–14 nm in diameter, are oriented perpendicularly to the cytoplasmic membrane and show a 6 nm repeat along their length. Isolated phycobilisomes of G. violaceus appear, after fixation and negative staining, as bundles of 6 parallel rodshaped elements connected to an ill-defined basal structure. The bundles are 40–45 nm wide and 75–90 nm long. The rods are 10–12 nm in width; their length varies between 50 and 70 nm. These rods are morphologically similar to those observed at the periphery of hemidiscoidal phycobilisomes of other cyanobacteria, with a strong repeat at 6 nm intervals and a weaker one at 3 nm intervals along their length. The calculated molar ratio of phycobiliproteins in isolated G. violaceus phycobilisomes corresponds to 1:3.9:2.9 for allophycocyanin, phycocyanin and phycoerythrin respectively. When excited at 500 nm, isolated phycobilisomes exhibit a major fluorescence emission band centered at 663 nm.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00425248
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  • 8
    Electronic Resource
    Electronic Resource
    Cloning and expression of a Nostoc sp. leucine biosynthetic gene in Escherichia coli (1986)
    Springer
    Archives of microbiology 145 (1986), S. 315-321 
    Details
    ISSN: 1432-072X
    Keywords: Cyanobacteria ; DNA restriction and cloning ; Gene fusions ; Leucine biosynthesis ; Mutant complementation ; Nostoc ; Transcription
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Genomic DNA extracted from the symbiotically-competent, heterocyst-forming cyanobacterium Nostoc sp. strain 7801 was resistant to cleavage by a number of restriction endonucleases. A cosmid library of Nostoc DNA was prepared and maintained in the modification-limited Escherichia coli strain HB101. Analysis of cloned Nostoc DNA fragments indicated infrequent occurrence of restriction endonuclease recognition sites in the Nostoc genome. The Nostoc genomic library was screened for sequences complementing mutations in the E. coli leucine and proline biosynthetic operons. Two cosmids complementing leuB were isolated but none for leuA, leuC, leuD, or proA were detected in 1000 cosmids. A 3.0 kb fragment subcloned from one of the cosmids complemented mutations in leuB when inserted into the HindIII site of pBR322 in either orientation, demonstrating that transcription of leuB originated within the cloned fragment. The cloned fragment also carries a second site capable of initiating transcription of fused antibiotic resistance genes. While transcription of Nostoc DNA sequences did occur in E. coli, unknown barriers must also exist that prevented additional biological complementation of specific E. coli mutations.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00470864
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  • 9
    Electronic Resource
    Electronic Resource
    The structure of cyanobacterial phycobilisomes: a model (1979)
    Springer
    Archives of microbiology 123 (1979), S. 113-127 
    Details
    ISSN: 1432-072X
    Keywords: Phycobilisomes ; Phycobiliproteins ; Cyanobacteria ; Chromatic adaptation ; Fine structure ; Photosynthesis ; Protein assembly
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Phycobilisomes, supramolecular complexes of water-soluble accessory pigments, serve as the major light-harvesting antennae in cyanobacteria and red algae. Regular arrays of these organelles are found on the surface of the thylakoid membranes of these organisms. In the present study, the hemi-discoidal phycobilisomes of several species of cyanobacteria were examined in thin sections of cells and by negative staining after isolation and fixation. Their fundamental structures were found to be the same. Isolated phycobilisomes possessed a triangular core assembled from three stacks of disc-shaped subunits. Each stack contained two discs which were ∼12 nm in diameter and ∼6–7 nm thick. Each of these discs was probably subdivided into halves ∼3–3.5 nm thick. Radiating from each of two sides of the triangular core were three rods ∼12 nm in diameter. Each rod consisted of stacks of 2 to 6 disc-shaped subunits ∼6 nm thick. These discs were subdivided into halves ∼3 nm thick. The average number of discs of ∼6 nm thickness forming the peripheral rods varied among the strains studied. For certain chromatically adapting strains, the average rod length was dependent upon the wavelength of light to which cells were exposed during growth. Analyses of phycobilisomes by spectroscopic techniques, polyacrylamide gel electrophoresis, and electron microscopy were compared. These analyses suggested that the triangular core was composed of allophycocyanin and that the peripheral rods contained phycocyanin and phycoerythrin (when present). A detailed model of the hemi-discoidal phycobilisome is proposed. This model can account for many aspects of phycobiliprotein assembly and energy transfer.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00446810
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  • 10
    Electronic Resource
    Electronic Resource
    Duplications created by transformation in Sordaria macrospora are not inactivated during meiosis (1989)
    Springer
    Molecular genetics and genomics 218 (1989), S. 390-396 
    Details
    ISSN: 1617-4623
    Keywords: Sordaria macrospora ; Transformation ; Filamentous fungi ; Orotate phosphoribosyl transferase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We present here the first report of a transformation system developed for the filamentous fungus Sordaria macrospora. Protoplasts from a ura-5 strain were transformed using the cloned Sordaria gene at a frequency of 2×10-5 transformants per viable protoplast (10 per microgram of DNA). Transformation occurred by integration of the donor sequences in the chromosomes of the recipient strain. In 71 cases out of 74, integration occurred outside the ura5 locus; frequently several (two to four) copies were found at a unique integration site. Using the advantage of the spore colour phenotype of the ura5-1 marker, we have shown that the transformed phenotype is stable through mitosis and meiosis in all transformants analysed. No methylation of the duplicated sequences could be observed during meiotic divisions in the transformants.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00332400
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