Figure 6

Physical mapping of Beta vulgaris chromoviruses. Localization of different chromoviruses on metaphase chromosomes and interphase nucleus of B. vulgaris by fluorescence in situ hybridization (FISH) using long terminal repeat (LTR)-specific probes. High-resolution FISH was performed on pachytene chromosomes and DNA fibers. In each panel, the stained (4^′,6^′-diamidino-2-phenylindole; DAPI: blue fluorescence) DNA shows the morphology of the chromosomes. Hybridized probes were detected with cyanine 3 (Cy3; red fluorescence) and fluorescein isothiocyanate (FITC; green fluorescence). (A) The LTR of Beetle 7 hybridized to centromeric and peri centromeric heterochromatin of most chromosomes; examples for depletion from centromeric heterochromatin are marked by arrows. (B) The hybridization with the LTR probe of Bongo 3 visualized the clustered organization of the Bongo 3 family along all chromosomes. Reduced hybridization to some centromeric regions was detectable (examples marked by arrows). (C) Amplification of Bingo 1 chromoviruses was seen as strong signals largely in the peri-centromeric regions. (D) The LTR of Beon 1 (red fluorescence) hybridized only to chromosome 1, harboring the ribosomal (r)RNA genes. (E-G) The localization of Beon 1 within the rDNA locus was confirmed by multi-color FISH analysis using a probe for the 18S rRNA (green fluorescence). (E) The signals of both DNA probes overlapped at the rDNA of the interphase nuclei. (F) FISH to chromosomes of B. vulgaris at the pachytene stage of meiosis showed interspersion of 18 rRNA genes with Beon 1 copies. (G) The hybridization to extended DNA fibers revealed the physical order of Beon 1 copies within the nucleolar organizer region (NOR) , where not every rDNA array was found to harbor a Beon 1 element.