Figure 1
Molecular cytogenetic and experimental breeding evidence for the ancestry of white clover from a hybrid between two species with contrasting habitats. (a, b) Habitat surroundings of T. occidentale (sea-level, SE Ireland) and T. pallescens (2,000m, Pyrenees, NE Spain), respectively. (c–e) Reciprocal GISH-FISH on an early metaphase cell from T. repens. (c) DAPI stained cell in grey-scale. Dotted guide-lines represent decondensed NORs. (d) GISH-FISH on the same cell as in c using genomic DNA of T. pallescens (green) and 18S rDNA (red). (e)Reprobing of the same cell as in d with genomic DNA of T. occidentale (red) and 5 S rDNA (green). Arrows indicate a T. occidentale-derived NOR chromosome pair and arrowheads a T. pallescens-derived chromosome pair with 5 S rDNA signals. (f) F1 hybrid plant 440–1. (gi) An early somatic metaphase cell from hybrid 440–1 subjected to reciprocal GISH-FISH. (g) DAPI stained cell in grey-scale. Dotted guideline represents a decondensed NOR.(h) GISH-FISH on the same cell as in g using genomic DNA of T. pallescens (green) and 18S rDNA (red). Arrow and arrowhead indicate T. occidentale- and T. pallescens-derived NOR chromosomes, respectively. (i) Reprobing of the same cell as in h with genomic DNA of T. occidentale (red) and 5S rDNA (green). Arrow and arrowhead indicate T. occidentale- and T. pallescens-derived chromosomes with 5S rDNA signals, respectively. (j–k) A meiotic metaphase cell from hybrid 440–1 subjected to GISH-FISH. (j) DAPI stained meiotic metaphase I cell in grey scale showing eight bivalents. (k) GISH-FISH on the same cell as in j using genomic DNA of T. pallescens (green) and 18S rDNA (red). Bivalent formation involving NOR chromosomes (arrow) derived from two parental species and differential hybridization of T. pallescens DNA indicate homoeologous pairing. Yellow colour of NORs in d, h, and k is due to simultaneous hybridization of T. pallescens DNA (green) and 18S rDNA (red). Bar (k) represents 10m and applies to all chromosomal micrographs.