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  • 1
    Electronic Resource
    Electronic Resource
    Male-specific DNA markers from African catfish (Clarias gariepinus) (2000)
    Springer
    Genetica 110 (2000), S. 267-276 
    Details
    ISSN: 1573-6857
    Keywords: Clarias ; RAPD ; sex determination ; siluroid ; teleost
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We searched for sex-specific DNA sequences in the male and female genomes of African catfish, Clarias gariepinus (Burchell, 1822) by comparative random amplified polymorphic DNA (RAPD) assays performed on pooled DNA samples. Two sex-linked RAPD markers were identified from the male DNA pool and confirmed on individual samples, showing good agreement with phenotypic sex. Both markers were isolated, cloned and characterized. The first marker (CgaY1) was nearly 2.6 kb long, while the length of second one (CgaY2) was 458 bp. Southern blot analysis with a CgaY1 probe showed strong hybridizing fragments only in males and not in females under stringent conditions, indicating the presence of multiple copies of CgaY1 in the male genome. When tested by zoo blot on the genomes of two closely related species from the Clariidae family, CgaY1 hybridized to the DNA of Heterobranchus longifilis and generated a faint male-specific band at low stringency. CgaY2 produced similar hybridization pattern in both sexes of C. gariepinus, C. macrocephalus and H. longifilis. Specific primers were designed to the sequences and the markers were amplified in multiplex PCR reactions together with a control band common to all individuals. This allowed for rapid, molecular sexing of the species on the basis of a simple three band (male) versus one band (female) pattern. According to our knowledge these are the first sex-specific DNA markers isolated from a siluroid fish species.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1012739318941
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  • 2
    Electronic Resource
    Electronic Resource
    Liposome-mediated gene transfer in fish embryos (1994)
    Springer
    Transgenic research 3 (1994), S. 116-119 
    Details
    ISSN: 1573-9368
    Keywords: reporter gene ; zygote ; transient expression ; bacteriophage lambda
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Liposome-mediated gene transfer was used to introduce large DNA constructs into zygotes of African catfish. The technique is based on the delivery of recombinant bacteriophage lambda particles (or DNA-protamine complexes) into the cytoplasm of target cells by negatively charged, large unilamellar liposomes. Dechorionated zygotes and early fish embryos were treated with the transforming liposomes. Expression of the introduced reporter genes during the first three weeks of the development of the larvae was followed by measuring the activity of corresponding enzymes. These assays have indicated very efficient DNA uptake into the embryos.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF01974089
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  • 3
    Electronic Resource
    Electronic Resource
    High transgene activity in the yolk syncytial layer affects quantitative transient expression assays in zebrafish (Danio rerio) embryos (1996)
    Springer
    Transgenic research 5 (1996), S. 433-442 
    Details
    ISSN: 1573-9368
    Keywords: zebrafish ; β-actin ; transient expression ; yolk syncytial layer ; polyploidy
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract For the porpose of studying the factors that cause wide variation in transient transgene expression in individual fish, alacZ reporter gene linked to a carp β-actin regulatory sequence was introduced into zebrafish embryos. As a general trend, a correlation between the number of transgene copies injected and the level of transgene expression was found. However, a substantial variation in the level of expression still occurred that could not be attributed to technical factors such as the difference in injected volume of the transgene. Co-injection of32P-dCTP and transgene into the same embryo followed by detection of β-galactosidase activity, has shown that the volume used for transgene injection, which was determined in terms of radioactivity, is not closely related to the level and location of transgene expression. Injection into the animal pole at zygote stage and the yolk cytoplasmic layer (YCL) at the 64-cell stage followed by determination of transgene expression in terms of unit injection volume, revealed that there are marked differences among tissues with regard to their capacity for transgene expression, and that the yolk syncytial layer is higher in this capacity. This high activity is assumed to be due to the high transcriptional activity or enhanced transgene replication in the syncytial layer, which is known to contain giant polyploid nuclei. The high level of expression in the YSL may influence transient expression studies using quantitative comparative analyses and should be taken into consideration when expression data are derived from homogenates of yolk sac embryos.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF01980208
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  • 4
    Electronic Resource
    Electronic Resource
    Agarose electrophoresis of DNA in discontinuous buffers, using a horizontal slab apparatus and a buffer system with improved properties (1993)
    Weinheim : Wiley-Blackwell
    Electrophoresis 14 (1993), S. 179-184 
    Details
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Using a horizontal slab apparatus with a buffer in the reservoirs at the level of the gel (“sea-level electrophoresis”), the retrograde discontinuous buffer system reported by Wiltfang et al. for sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of proteins was applied to DNA electrophoresis. This application yielded the advantages of an increased displacement rate of the moving boundary front and a decrease in the concentration of the counterion base in the resolving phase, which yielded reduced relative mobility values at equivalent gel concentrations and practicable low buffer concentrations. The change of relative mobilities (Rf) with a variation of field strength is decreased compared to that of the migration rate in the continuous Tris-boric-acid-EDTA (TBE) buffer and thus the robustness of the system is improved, as well as the efficiency of separation. The system of Wiltfang et al. has in common with previously described discontinuous DNA systems, that it is able to stack DNA from dilute samples and is insensitive to sample components with lower net mobilities than DNA, such as acetate. However, the variance of Rf at constant current density in the discontinuous buffer system is not improved over that of the migration rate at constant field strength in the continuous TBE buffer.
    Additional Material: 7 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150140130
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  • 5
    Electronic Resource
    Electronic Resource
    An improved voltage measurement device for gel electrophoresis in tube apparatus (1988)
    Weinheim : Wiley-Blackwell
    Electrophoresis 9 (1988), S. 32-36 
    Details
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: A device for the measurement of voltage across tube gels was designed and constructed which allows one to measure voltage during electrophoresis without any manipulation of the gel electrophoresis apparatus or gel tube and with the elimination of a source of inaccuracy in previous such devices.
    Additional Material: 4 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150090107
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  • 6
    Electronic Resource
    Electronic Resource
    Physical identification of a virus in a crude leaf extract by its ferguson plot in agarose gel electrophoresis (1988)
    Weinheim : Wiley-Blackwell
    Electrophoresis 9 (1988), S. 162-166 
    Details
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Crude extracts of turnip crinkle virus upon agarose gel electrophoresis yield (i) virus patterns unperturbed by contaminants; (ii) plots of mobility vs. gel concentration (Ferguson plots) parallel with those of the purified virus. The parallelism suggests similarity in size and shape but a lower net charge for the crude virus. This result is obtained when gel electrophoresis is carried out either in a continuous buffer or in a discontinuous (moving boundary electrophoresis) buffer system. The latter mode has the substantial benefit of electrophoretic (auto-)concentration of dilute virus sample prior to resolution. Thus, the Ferguson plot analysis in a discontinuous buffer system of turnip crinkle virus can be viewed as a model procedure for the physical identification of other viruses contained in dilute extracts, feasible even in the absence of a prior knowledge as to the nature of, or isolation of, the virus.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150090403
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  • 7
    Electronic Resource
    Electronic Resource
    Sieving of ionic constituents across moving boundaries in gel electrophoresis (1989)
    Weinheim : Wiley-Blackwell
    Electrophoresis 10 (1989), S. 254-259 
    Details
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: The representative β-hydroxyethylmorpholinium-chloride-bicinate moving boundary with a trailing ion net mobility relative to Na+ of 0.41, detected by precipitation of chloride with silver nitrate, exhibits a decreasing chloride mobility at increasing polyacrylamide gel concentrations from 3.5 to 45 %T, 5 %CBis. This decrease, largely due to an increase of field strength at constant current, is described by a convex plot of log (mobility) vs. %T (Ferguson plot) and signifies that chloride/bicinate are sieved by the gel. In agarose gels, the same plot of mobility vs. gel concentration is constant below 7 % gel concentration, since in those gels field strength and migration rate remain the same within that gel concentration range. Both in polyacrylamide and in agarose gels the displacement rate of the chloride-bicinate boundary as a function of the time of electrophoresis or distance migrated remains invariant within 15 %. The plot of log(mobility) vs. gel concentration extrapolated to 0 %T is 5.85 and 5.41 (10-5 cm2 s-1V-1) for polyacrylamide and for agarose (SeaKem HGT-P, FMC) gels, respectively. The slightly decreased mobility intercept at 0 %T for agarose is presumably due either to the electroendosmotic properties of agarose HGT-P and/or failure to Sufficiently take into account the flattening of the Ferguson plot in the polyacrylamide concentration range below 3 % in which a transition from a gel to a fluid (sol) medium takes place.
    Additional Material: 2 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150100407
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  • 8
    Electronic Resource
    Electronic Resource
    A thin-layer multistrip polyacrylamide gel electrophoresis apparatus for ferguson plot analysis at the sub-microgram load level (1989)
    Weinheim : Wiley-Blackwell
    Electrophoresis 10 (1989), S. 726-729 
    Details
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: A procedure was developed for casting thin-layer multistrip polyacrylamide gels and using them for the simultaneous gel electrophoresis at several gel concentrations (Ferguson plot analysis) at the sub-microgram load level, using silver staining, autoradiography and, potentially, blotting for detection. The lower viscosity of polymerization mixtures, compared to agarose gelation mixtures, required the redesign of the multistrip cassette with separation of channels by rubber gaskets and the application of a cassette press. The lowered viscosity also required addition of 35% sucrose and an increased rate of polymerization in application to multistrip gels formed on a common NetFix backing. The present design allows one to obtain Ferguson plots exemplified by those of 32P-labeled DNA followed by autoradiographic detection.
    Additional Material: 4 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150101013
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  • 9
    Electronic Resource
    Electronic Resource
    Detection of turnip crinkle virus on agarose gel electropherograms at the nanogram load level (1988)
    Weinheim : Wiley-Blackwell
    Electrophoresis 9 (1988), S. 299-302 
    Details
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: The previous conditions for the physical characterization of turnip crinkle virus (TCV) by quantitative agarose gel electrophoresis [1, 2] were limiting the method to the microgram load level and were therefore insufficiently sensitive to satisfy the need in many areas of virology for detection of viruses containing single-stranded RNA at the nanogram level. The present report remedies that defect by presenting a technique compatible with the nanogram load level of such viruses. The technique is based on a reduction of gel thickness and on the use of silver staining.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150090703
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  • 10
    Electronic Resource
    Electronic Resource
    Characterization of the electrophoretic properties of nucleosome core particles by transverse polyacrylamide pore gradient gel electrophoresis (1993)
    Weinheim : Wiley-Blackwell
    Electrophoresis 14 (1993), S. 720-724 
    Details
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Transverse pore gradient gel electrophoresis, previously applied to bent DNA, has extended the usefulness of the gel retardation assay in two ways: (i) by differentiating between different DNA conformations; (ii) by providing information regarding the physical properties of DNA. In the present study, similarly extended information is obtained with regard to a well-characterized DNA-protein complex, the chicken erythrocyte nucleosome core particle. (i) The winding of DNA around the protein core constrains the DNA which renders its Ferguson curve (migration distance vs. gel concentration) similar to that of kinetoplast DNA, i.e. it intersects sharply with the Ferguson curves of linear DNA standards. By contrast, the deproteinized nucleosome DNA exhibits a Ferguson curve similar to linear standards of the same lenght. (ii) Interpretation of the Ferguson curve based on a mathematical model shows that the nucleosome exhibits a linear Ferguson plot [log(mobility) vs. gel concentration]. This is similar to and characteristic of spherical proteins, contrasting with the concave plot typical for linear and bent DNA. (iii) The effective size of the nucleosome, evaluated in terms of an “equivalent sphere” (i.e. a hypothetical spherical particle with a radius, Res, having the same electrophoretic mobility as DNA for a particular set of experimental conditions), remains invariant across the gel concentration range of 3-9 % T. This is similar to proteins and bacteriophages and contrasts with the progressive decline of Res with increasing gel concentration observed for linear DNA and the deproteinized nucleosomal DNA.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.11501401114
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