Abstract
With the advent of nonradioactive labels, in situ hybridization (ISH) has become a useful technique for the detection of viral DNA in infected tissue (1), mRNA expression (2), sex determination (3), human gene mapping (4), and interphase cytogenetics (5–8). For chromosomal mapping of genes by ISH, labeled DNA probe is hybridized to metaphase spreads. If the label is a radioisotope, the signal is detected by autoradiography (9). Nonradioactive labels, such as biotin (4), digoxigenin (10), or 2-acetylaminofluorene (AAF), are detected by immunocytochemistry. AAF is incorporated by chemical modification of the DNA probe. Biotin and digoxigenin are incorporated enzymatically by nick translation or by random primer extension (11), although both can be incorporated by chemical modification (12). In the authors’ laboratory, biotin has been successfully used in mapping genes with probes of 0.8 Kb (10). The method described here, therefore, applies to biotin-labeled probes although, with minor modifications, can be adapted to probes labeled by digoxigenin or AAF. An example of chromosomal mapping of a unique sequence is shown in Fig. 1.
Human chromosome spread probed with a biotdnylated β-globin probe. There is a signal at band pl.5 (arrow) on chromosome 11.
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Further Reading
Polak, J. M. and McGee, J. O’D, eds. (1990) In Situ Hybridisation: Principles and Practice. Oxford University Press, UK.
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Bhatt, B., McGee, J.O. (1992). Chromosomal Mapping of Genes by Nonisotopic In Situ Hybridization. In: Manson, M.M. (eds) Immunochemical Protocols. Methods in Molecular Biology, vol 10. Humana Press, Totowa, NJ. https://doi.org/10.1385/0-89603-204-3:421
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DOI: https://doi.org/10.1385/0-89603-204-3:421
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