Supplementary Material for: Molecular Cytogenetics (FISH, GISH) of <b><i>Coccinia grandis</i></b>: A ca. 3 myr-Old Species of Cucurbitaceae with the Largest Y/Autosome Divergence in Flowering Plants

The independent evolution of heteromorphic sex chromosomes in 19 species from 4 families of flowering plants permits studying X/Y divergence after the initial recombination suppression. Here, we document autosome/Y divergence in the tropical Cucurbitaceae <i>Coccinia grandis</i>, which is ca. 3 myr old. Karyotyping and C-value measurements show that the <i>C. grandis</i> Y chromosome has twice the size of any of the other chromosomes, with a male/female C-value difference of 0.094 pg or 10% of the total genome. FISH staining revealed 5S and 45S rDNA sites on autosomes but not on the Y chromosome, making it unlikely that rDNA contributed to the elongation of the Y chromosome; recent end-to-end fusion also seems unlikely given the lack of interstitial telomeric signals. GISH with different concentrations of female blocking DNA detected a possible pseudo-autosomal region on the Y chromosome, and C-banding suggests that the entire Y chromosome in <i>C. grandis</i> is heterochromatic. During meiosis, there is an end-to-end connection between the X and the Y chromosome, but the X does not otherwise differ from the remaining chromosomes. These findings and a review of plants with heteromorphic sex chromosomes reveal no relationship between species age and degree of sex chromosome dimorphism. Its relatively small genome size (0.943 pg/2C in males), large Y chromosome, and phylogenetic proximity to the fully sequenced <i>Cucumis sativus</i> make <i>C. grandis</i> a promising model to study sex chromosome evolution