We have previously developed the technology to identify active gene regions on animal cell metaphase chromosomes. This method (D-banding) is based on the sensitivity of active genes for the enzyme DNaseI. Chromosome preparations are nick translated in situ using DNaseI and DNA polymerase and the substituted active regions visualized by autoradiography or by biotin detection. We will use this methodology to study the relationship between gene transcription and DNA replication and to search for tissue specific chromosomal domains. Cell-type specific gene clusters will also be analyzed by in situ hybridization using probes from a CHO liver specific library which we are preparing. We will use the D-banding technology to study the mechanism of regulation of domains of genes as is seen in phenotypic switching and allelic silencing. Finally, we will study the molecular basis of X-chromosome activation and inactivation, using 5-azacytidine treatment to induce changes in the activity state of this chromosome in various cell types in culture. These experiments should pave the way for the cloning of the gene for a cellular factor which interacts with the X chromosome and causes its activation. The methodologies allow, for the first time, the study of chromosome gene structure and function at the morphological level.
| Jablonka, E; Goitein, R; Sperling, K et al. (1987) 5-aza-C-induced changes in the time of replication of the X chromosomes of Microtus agrestis are followed by non-random reversion to a late pattern of replication. Chromosoma 95:81-8 |
| Jablonka, E; Goitein, R; Marcus, M et al. (1985) DNA hypomethylation causes an increase in DNase-I sensitivity and an advance in the time of replication of the entire inactive X chromosome. Chromosoma 93:152-6 |
