Direct FISH of 5S rDNA on tomato pachytene chromosomes places the gene at the heterochromatic knob immediately adjacent to the centromere of chromosome 1.

We describe a molecular cytogenetic procedure for high resolution physical mapping of DNA markers, an essential step toward construction of an integrated molecular-classical-cytological map. Tomato was selected as material because its pachytene chromosomes are amenable for study and because detailed molecular, classical, and cytological maps are available. Karyotyping of acetocarmine-stained pachytene chromosomes showing detailed cytogenetic landmarks was combined with direct FISH of the 5S rDNA gene. This enabled us to pinpoint the 5S rDNA gene to the first heterochromatic knob immediately adjacent to the centromere in the short arm of chromosome 1. Thus the position of the 5S rDNA gene on the molecular map was related to the position of the 5S rDNA on the cytogenetic map. The results also provide conclusive evidence of the location of a functional gene in the pericentric heterochromatic region, a rare event to date in plants. We conclude that karyotyping of pachytene chromosomes can be combined with FISH to map a DNA sequence to a cytogenetically defined region and to determine the chromatin origin of an expressed gene. Key words : direct fluorescence in situ hybridization, 5S rDNA, pachytene chromosomes, heterochromatic gene.