Stability of DNA content and molecular cytogenetic markers in the genus Stylosanthes (Fabaceae)

Abstract

Stylosanthes (Leguminosae) comprises about 50 species being some of them with great economic importance. Some species are widely used as forages, to recover degraded areas due to its adaptability to low fertility soils and nitrogen fixing capacity as well as a green fertilizer. The genus is distributed in tropical and subtropicals areas. Brazil is considered the larger diversity center, with great inter and intraspecific variation. Breeding studies have increased, however, the taxonomic situation of the group remains unsettled and controversial. The occurrence of polyploidy, hybrids and many additional synonyms reinforce the importance to add new information about the genus. Cytogenetic data are scarce, even considering as an important tool to give relevant information about the genome. Here we describe, for the first time, karyotypic aspects and the DNA content of diploid species (S. grandifolia, S. gracilis, S. hipocampoides, S. acuminata and four varieties of S. guianensis) by using molecular cytogenetic techniques (CMA/DAPI banding and in situ hybridization) and flow cytometry. Seeds germination was induced by temperature variation (18° to 40°C) and growth hormones (CEPA 0.1mM) in a germination chamber. Meristematic roots were used to slides preparation. Chromosomes were stained with CMA/DAPI for banding patterns analysis and ribosomal 5S, 45S rDNA sites were mapped by in situ hybridization technique. To estimate the DNA content, Pisum sativum were used as a reference standard. The chromosomal morphometry showed a symmetrical karyotype with few variations among the species. The number of CG rich regions ranged from 2 to 4. Regarding 5S and 45S rDNA, two sites were observed in most species, excepting in S. acuminata, in which four 45S markers were identified. DNA content estimation was about 2.50pg in all species. These new data constitutes an important contribution in the study of evolutionary history of the group, opening new possibilities for future comparisons among genomes of different Stylosanthes species and also being of great value for breeding programs of species with economic importance.