6th ESACP Congress, Heidelberg, April 7-11, 1999

A049
REPRODUCIBLE FOLDING OF AN AMPLIFIED DNA SEQUENCE TO THE METAPHASE CHROMATID AXIS
Dietzel S, Belmont AS

University of Illinois at Urbana-Champaign, Cell and Structural Biology, Urbana, Illinois, USA

A cell clone displaying a localization of an amplified sequence to the axis of metaphase and early anaphase chromatids is described. The amplified, GFP labeled sequences form a line along the axis of the daughter chromatids, with the measured diameter of the GFP-signal close to the resolution limit of the light microscope and roughly ¢ the chromatid diameter. The gene amplified chromosome region contains vector insertion sites with lac operator repeats which were detected by a lac-repressor-GFP fusion protein (Robinett et al., 1996, J. Cell Biol.135:1685; Li et al., 1998, J. Cell Biol. 140:975). Within interphase nuclei, these integration sites can be detected as a number of GFP-fluorescent dots, restricted to one or two areas in the nucleus, corresponding to individual chromosomes carrying the sequences. The tagged sequences are located at the periphery of condensing prophase chromatids and do not move to the axial position until the prophase/metaphase transition. Live cell observation showed that the linear alignment is lost early in G1. Our results will be compared to existing models of mitotic chromosome organization.