The spatial localization of T-DNA insertions in petunia interphase nuclei: consequences for chromosome organization and transgene insertion sites.

In an earlier fluorescent in-situ hybridization (FISH) study on petunia (ten Hoopen et al. 1996), we found a considerable discrepancy between the genetic map and the physical map with respect to T-DNA insertions on metaphase chromosomes. For some transgenes we found a preference to integrate near the telomeres. Here, we studied the spatial position of transgenes in interphase nuclei by FISH and 3D-confocal microscopy to elucidate a possible structural preference for the nuclear localization of transgenes. Three transgenes located near telomeres on three different metaphase chromosomes showed a much more internal distribution in interphase root meristem than the telomeres, whereas a proximal transgene appeared to be distributed in a random fashion. The results point to local differences in chromatin compacting along a chromosome. These differences might explain a preference for T-DNA insertion in distal regions of the chromosome.

Localization of T-DNA Insertions in Petunia by Fluorescence in Situ Hybridization: Physical Evidence for Suppression of Recombination.

Using fluorescence in situ hybridization (FISH) with metaphase preparations, we localized a 4-kb single-copy T-DNA sequence in a group of petunia transformants. The selected T-DNAs previously had been shown to be linked to the phenotypic marker FI on chromosome II. Linkage analysis had revealed that recombination around the FI locus is suppressed in a wide cross relative to an inbred recombination assay. The localization of six FI-linked T-DNAs and the FI locus itself, using FISH, revealed a number of aspects of recombination in petunia: (1) the central region of chromosome II showed at least a 10-fold suppression of recombination in wide crosses relative to the distal region; (2) recombination in wide hybrids over two-thirds of the chromosome was extremely low; and (3) recombination between completely homologous chromosomes in an inbred cross also was suppressed in the central region. In addition, the T-DNAs were not evenly distributed along the chromosome, suggesting a possible preference for a distal position for T-DNA integration. Implications for such a preference are discussed.