Chromosomal location of endogenous geminivirus-related DNA sequences in Nicotiana tabacum L

The N. tabacum (tobacco) nuclear genome carries approximately 25 multiple direct repeats of a geminivirus-related DNA (GRD) sequence that probably arose by illegitimate recombination, following geminivirus infection, during Nicotiana evolution. Each GRD repeat carries sequences similar to the geminiviral AL1 gene of the tomato golden mosaic virus (TGMV), encoding a protein required for viral DNA replication, plus the cis-essential replication origin. Using a cloned 14-kb GRD repeat sequence as a probe for fluorescence in situ hybridization (FISH), we identified a unique tobacco chromosome carrying GRD. Translocations between chromosomes of the tobacco S and T genomes were used as physical markers by sequentially hybridizing chromosomes with labelled GRD and total genomic DNA from N. sylvestris (equivalent to the S genome). The 25S, 18S and 5.8S ribosomal gene clusters were detected in double-labelling experiments for use as additional markers to identify the chromosomal location of GRD. GRD occupies one site on a homologous pair of small submetacentrics from the T genome characterized by a lack of either translocated segments from the S genome or ribosomal genes. GRD provides an additional marker for the small chromosomes of the T genome and a useful phylogenetic tool.