Mapping ESTs by Fiber–FISH
- Nina Horelli-Kuitunen1,6,
- Johanna Aaltonen2,
- Marie-Laure Yaspo3,
- Mervi Eeva1,
- Maija Wessman1,4,
- Leena Peltonen2,5, and
- Aarno Palotie1,5
- 1Departments of Clinical Chemistry and Biomedicine, University of Helsinki and Laboratory, Department of Helsinki University Central Hospital, 00290 Helsinki, Finland; 2National Public Health Institute, Department of Human Molecular Genetics, 00300 Helsinki, Finland and Haartman Institute, Department of Medical Genetics, University of Helsinki, 00014 Helsinki, Finland; 3 Max Planck Institute for Molecular Genetics, D-14195 Berlin, Germany; 4 Department of Biosciences, Division of Genetics, University of Helsinki, 00014 Helsinki, Finland
Abstract
A visual transcript map of six genes was constructed on the chromosome 21q22.3 by high resolution fluorescence in situ hybridization (FISH). Expressed sequence tags (ESTs) from six genes—PWP2, KNP1, AIRE, C21orf3,SMT3A, and C21orf1—were successfully localized by fiber–FISH by use of sensitive tyramide-based detection. The sizes of the ESTs varied between 315 to 956 bp and most of them map within the 3′-untranslated region. The ESTs were assigned to and subsequently ordered within cosmid, PAC, and BAC clones hybridized on DNA fibers. Physical distances between ESTs and known markers were determined. Our results demonstrate the feasibility and accuracy of visual mapping EST sequences in relation to known markers. The main advantage of this approach is that it can be applied to finely map any of the database ESTs for positional cloning efforts. The sensitivity, specificity, and reproducibility of this high-resolution EST mapping technique is evaluated.
Footnotes
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↵5 Present address: Department of Human Genetics and Department of Pathology, University of California Los Angeles, Los Angeles, California 90095-7088 USA.
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↵6 Corresponding author.
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E-MAIL Nina.Horelli-Kuitunen{at}HUCH.fi; FAX 358-9-4714001.
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- Received June 30, 1998.
- Accepted November 16, 1998.
- Cold Spring Harbor Laboratory Press
