Library

X
feed icon rss

Your email was sent successfully. Check your inbox.

An error occurred while sending the email. Please try again.

Proceed reservation?

Export
  • 1
    Electronic Resource
    Electronic Resource
    Marker pattern instabilities as a major cause of reproducibility problems in two-dimensional DNA fingerprinting (1996)
    Weinheim : Wiley-Blackwell
    Electrophoresis 17 (1996), S. 659-666 
    Details
    ISSN: 0173-0835
    Keywords: Two-dimensional DNA fingerprinting ; Denaturing gradient gel electrophoresis ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Two-dimensional (2-D) DNA fingerprinting is a promising technique for multilocus analysis of eukaryotic genomes. It has been successfully applied to the detection of DNA variation in tumors, to linkage analyses and to genomic comparisons of inbred mouse strains. However, there are still problems with inter-gel comparisons of 2-D DNA typing patterns as documented by the intergel reproducibility rates reported in the literature, which range from 84 to 98%. The basis for standardization in almost all of these studies has been a set of lambda fragments (digested separately with the restriction enzymes HaeIII, RsaI, Bg/I) that produces a spot pattern scattered across the gel. These spots are used as markers for gel comparisons. Since we noticed considerable variations in the marker spot paterns, we evaluated the properties of the lambda marker using both computer simulation and an empirical analysis of forty independent consecutive gels from our laboratory. We explain the instabilities of the spot pattern on the basis of the melting properties of the individual lambda fragments. A subset of spots is presented that has been stable in all our experiments. Only this set of spots should be used for gel standardization purposes until a new, completely reproducible marker becomes available. Finally, suggestions for an improved marker system are made.
    Additional Material: 6 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150170406
    Library Location Call Number Volume/Issue/Year Availability
    BibTip Others were also interested in ...
    Paper (German National Licenses)
    Fulltext
  • 2
    Electronic Resource
    Electronic Resource
    A novel standardization method for two-dimensional DNA fingerprints (1997)
    Weinheim : Wiley-Blackwell
    Electrophoresis 18 (1997), S. 2874-2879 
    Details
    ISSN: 0173-0835
    Keywords: DNA fingerprinting ; Two-dimensional DNA electrophoresis ; Numerical iteration ; Relaxation method ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Two-dimensional (2-D) DNA fingerprinting is a technique that allows for parallel genome analysis through the simultaneous detection of up to 500 minior microsatellite loci on a 2-D gel. Separation is performed according to size and melting temperature in the gel. In the application of this technique in genome analysis, a standardized method for the identification of individual spots is required. However, due to the polymorphic nature of up to 80% of the spots, existing standardization methods that have been primarily developed for 2-D protein patterns are not suitable for this task. We developed a robust method that standardizes 2-D DNA fingerprint spots on the basis of melting temperature - or denaturing gradient position - and fragment size. An external marker was used as a basis for standardization. A normalization surface was calculated over the gel dimensions by adapting an established numerical iteration technique previously used in physics termed “relaxation method”. The relaxation method works robustly with the irregularly spaced marker spots. The evaluation of the method for a spot of preknown position derived from the TP53 gene revealed a median observed error below 1% for fragment length and denaturing gradient position. The search for candidate minisatellite loci in genomic difference analysis depends on the reliable identification of alleles of this locus in different individuals. We proved experimentally that alleles of a single minisatellite locus cloned from a 2-D gel cluster on an isothermal line can be reliably identified using the presented standardization method. In conclusion, a standardization tool for a broader application of 2-D DNA fingerprinting in both tumor analysis and possibly parallel mutation screening is now available.
    Additional Material: 6 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150181527
    Library Location Call Number Volume/Issue/Year Availability
    BibTip Others were also interested in ...
    Paper (German National Licenses)
    Fulltext
  • 3
    Electronic Resource
    Electronic Resource
    Parallel genome analysis by one- and two-dimensional DNA fingerprinting in human gliomas (1995)
    Weinheim : Wiley-Blackwell
    Electrophoresis 16 (1995), S. 1715-1725 
    Details
    ISSN: 0173-0835
    Keywords: DNA fingerprinting ; Two-dimensional DNA typing ; Genome scanning ; Tumor analysis ; Gliomas ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: The detection of DNA variation in cancers is an important step in elucidating the mechanism of tumorigenesis. Using the strategy of multipoint genome analysis we detected many differences between glioma-derived and constitutional DNA by customary DNA fingerprinting with simple repetitive oligonucleotide probes. Amplification of the epidermal growth factor receptor (EGFR) gene has been found to be easily detectable as new or highly intensified bands in one-dimensional (1-D) DNA fingerprints of glioblastoma DNA generated with probes (GTG)5 or (GT)8. However, in most low-grade astrocytomas, 1-D DNA fingerprinting has failed to reveal any genomic abnormalities. In these cases a two-dimensional (2-D) technique was successfully employed that is based on size separation in neutral gels followed by sequence-dependent separation in denaturing gradient gels and hybridization with several mini- and microsatellite core probes. The hundreds of spots visualized with this technique were used to detect subtle changes probably occurring as the initial steps of tumorigenesis in human gliomas. On average, five of the approximately 580 sports generated by probes CAC and 33.6 were found to be altered in tumor DNA; 80% of the alterations were spot losses, the rest being spot gains or amplifications. Computer-based image analysis using an external lambda marker provided a stringent way to compare spot patterns generated by 2-D DNA finger-printing. In comparisons performed between typing patterns generated on the same gel, 99% of truly identical spots were confirmed by the sofware. In intergel comparisons 84% of identical spots were matched on the basis of the marker information alone.
    Additional Material: 7 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.11501601284
    Library Location Call Number Volume/Issue/Year Availability
    BibTip Others were also interested in ...
    Paper (German National Licenses)
    Fulltext
  • 4
    Electronic Resource
    Electronic Resource
    Cloning of minisatellite-containing sequences from two-dimensional DNA fingerprinting gels reveals the identity of genomic alterations in low-grade gliomas of different patients (1997)
    Weinheim : Wiley-Blackwell
    Electrophoresis 18 (1997), S. 1586-1591 
    Details
    ISSN: 0173-0835
    Keywords: Two-dimensional DNA fingerprinting ; Gliomas ; Genomic changes ; Spot cloning ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Two-dimensional (2-D) DNA fingerprinting was used to investigate genomic changes in human low-grade gliomas of different subtypes. DNA variations were identified in the 2-D hybridization patterns as spot losses or gains. Computer-aided matching of spot patterns from different patients revealed a clustering of spot changes at particular areas in the gel. Representative spots of each cluster were cloned using a spot cloning protocol which includes the preparation of a duplicate and a master gel. The DNA fragments of the 2-D gels were transferred to DEAE and nylon membrane, respectively. After hybridization of the master blot with a minisatellite core probe, the position of a particular spot was determined with reference to the lambda DNA fragments used as external markers in both gels. The gel spot DNA was recovered from the DEAE membrane by high salt elution and was polymerase chain reaction (PCR)-amplified after ligation of adaptor oligo cassettes. The PCR products were cloned and used as locus-specific probe for the rehybridization of the 2-D blots. One of these probes detected a spot loss in 7 of 28 low-grade gliomas of different subtypes analyzed. Another probe revealed a characteristic intensity shift in 8 of 9 pilocytic astrocytomas between two neighboring spots. The target sequence of this highly specific effect was assigned to chromosome 11q14 by in situ hybridization of a P1 clone harboring the affected genomic region. Thus, we successfully established a spot cloning procedure for the generation of locus-specific probes that may be instrumental in the discovery of the ciritical early events of glioma pathogenesis.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150180917
    Library Location Call Number Volume/Issue/Year Availability
    BibTip Others were also interested in ...
    Paper (German National Licenses)
    Fulltext
Close ⊗
This website uses cookies and the analysis tool Matomo. More information can be found here...