Chromosomal Mapping of Genes by Nonisotopic In Situ Hybridization
互联网
574
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 .
Fig. 1.
Human chromosome spread probed with a biotdnylated β-globin probe. There is a signal at band pl.5 (arrow) on chromosome 11.
