Gene mapping on human metaphase chromosomes by in situ hybridization with 3H, 35S, and 32P labeled probes and transmission electron microscopy

Abstract

A method based on in situ hybridization, autoradiography and transmission electron microscopy for mapping genes on human metaphase chromosomes is presented. Successful mapping of the tandemly repeated rDNA genes and of two nucleic acid probes, N-myc and probe 3 (Kanda et al. 1983), that are amplified in a homogeneously staining region (HSR) of the neuroblastoma cell line, IMR-32 is described. By using sufficiently thin AgBr emulsions, it is possible to obtain observable grains and good resolution with probes radiolabeled with 3H, 35S, or 32P, but the former gives the best results. We observe that neither of the two probes, N-myc and probe 3, has a uniform spatial distribution along the HSR and that the distributions of the two probes differ from each other. These observations support previous studies which indicated that the formation of an HSR is a more complex process than uniform amplification of a single DNA segment to form an n-fold set of perfect tandem repeats. The present study shows that the electron microscopic method is useful for extending the results of light microscopic studies for problems where higher resolution mapping is needed.