Cytomolecular diversity among Vigna Savi (Leguminosae) subgenera.

The genus Vigna (Leguminosae) comprises about 150 species grouped into five subgenera. The present study aimed to improve the understanding of karyotype diversity and evolution in Vigna, using new and previously published data through different cytogenetic and DNA content approaches. In the Vigna subgenera, we observed a random distribution of rDNA patterns. The 35S rDNA varied in position, from terminal to proximal, and in number, ranging from one (V. aconitifolia, V. subg. Ceratotropis) to seven pairs (V. unguiculata subsp. unguiculata, V. subg. Vigna). On the other hand, the number of 5S rDNA was conserved (one or two pairs), except for V. radiata (V. subg. Ceratotropis), which had three pairs. Genome size was relatively conserved within the genus, ranging from 1C = 0.43 to 0.70 pg in V. oblongifolia and V. unguiculata subsp. unguiculata, respectively, both belonging to V. subg. Vigna. However, we observed a positive correlation between DNA content and the number of 35S rDNA sites. In addition, data from chromosome-specific BAC-FISH suggest that the ancestral 35S rDNA locus is conserved on chromosome 6 within Vigna. Considering the rapid diversification in the number and position of rDNA sites, such conservation is surprising and suggests that additional sites may have spread out from this ancestral locus.

Translocations and inversions: major chromosomal rearrangements during Vigna (Leguminosae) evolution.

KEY MESSAGE: Inversions and translocations are the major chromosomal rearrangements involved in Vigna subgenera evolution, being Vigna vexillata the most divergent species. Centromeric repositioning seems to be frequent within the genus. Oligonucleotide-based fluorescence in situ hybridization (Oligo-FISH) provides a powerful chromosome identification system for inferring plant chromosomal evolution. Aiming to understand macrosynteny, chromosomal diversity, and the evolution of bean species from five Vigna subgenera, we constructed cytogenetic maps for eight taxa using oligo-FISH-based chromosome identification. We used oligopainting probes from chromosomes 2 and 3 of Phaseolus vulgaris L. and two barcode probes designed from V. unguiculata (L.) Walp. genome. Additionally, we analyzed genomic blocks among the Ancestral Phaseoleae Karyotype (APK), two V. unguiculata subspecies (V. subg. Vigna), and V. angularis (Willd.) Ohwi & Ohashi (V. subg. Ceratotropis). We observed macrosynteny for chromosomes 2, 3, 4, 6, 7, 8, 9, and 10 in all investigated taxa except for V. vexillata (L.) A. Rich (V. subg. Plectrotropis), in which only chromosomes 4, 7, and 9 were unambiguously identified. Collinearity breaks involved with chromosomes 2 and 3 were revealed. We identified minor differences in the painting pattern among the subgenera, in addition to multiple intra- and interblock inversions and intrachromosomal translocations. Other rearrangements included a pericentric inversion in chromosome 4 (V. subg. Vigna), a reciprocal translocation between chromosomes 1 and 5 (V. subg. Ceratotropis), a potential deletion in chromosome 11 of V. radiata (L.) Wilczek, as well as multiple intrablock inversions and centromere repositioning via genomic blocks. Our study allowed the visualization of karyotypic patterns in each subgenus, revealing important information for understanding intrageneric karyotypic evolution, and suggesting V. vexillata as the most karyotypically divergent species.

Oligo-FISH barcode in beans: a new chromosome identification system.

KEY MESSAGE: An Oligo-FISH barcode system was developed for two model legumes, allowing the identification of all cowpea and common bean chromosomes in a single FISH experiment, and revealing new chromosome rearrangements. The FISH barcode system emerges as an effective tool to understand the chromosome evolution of economically important legumes and their related species. Current status on plant cytogenetic and cytogenomic research has allowed the selection and design of oligo-specific probes to individually identify each chromosome of the karyotype in a target species. Here, we developed the first chromosome identification system for legumes based on oligo-FISH barcode probes. We selected conserved genomic regions between Vigna unguiculata (Vu, cowpea) and Phaseolus vulgaris (Pv, common bean) (diverged ~ 9.7-15 Mya), using cowpea as a reference, to produce a unique barcode pattern for each species. We combined our oligo-FISH barcode pattern with a set of previously developed FISH probes based on BACs and ribosomal DNA sequences. In addition, we integrated our FISH maps with genome sequence data. Based on this integrated analysis, we confirmed two translocation events (involving chromosomes 1, 5, and 8; and chromosomes 2 and 3) between both species. The application of the oligo-based probes allowed us to demonstrate the participation of chromosome 5 in the translocation complex for the first time. Additionally, we detailed a pericentric inversion on chromosome 4 and identified a new paracentric inversion on chromosome 10. We also detected centromere repositioning associated with chromosomes 2, 3, 5, 7, and 9, confirming previous results for chromosomes 2 and 3. This first barcode system for legumes can be applied for karyotyping other Phaseolinae species, especially non-model, orphan crop species lacking genomic assemblies and cytogenetic maps, expanding our understanding of the chromosome evolution and genome organization of this economically important legume group.

BAC- and oligo-FISH mapping reveals chromosome evolution among Vigna angularis, V. unguiculata, and Phaseolus vulgaris.

Cytogenomic resources have accelerated synteny and chromosome evolution studies in plant species, including legumes. Here, we established the first cytogenetic map of V. angularis (Va, subgenus Ceratotropis) and compared this new map with those of V. unguiculata (Vu, subgenus Vigna) and P. vulgaris (Pv) by BAC-FISH and oligopainting approaches. We mapped 19 Vu BACs and 35S rDNA probes to the 11 chromosome pairs of Va, Vu, and Pv. Vigna angularis shared a high degree of macrosynteny with Vu and Pv, with five conserved syntenic chromosomes. Additionally, we developed two oligo probes (Pv2 and Pv3) used to paint Vigna orthologous chromosomes. We confirmed two reciprocal translocations (chromosomes 2 and 3 and 1 and 8) that have occurred after the Vigna and Phaseolus divergence (~9.7 Mya). Besides, two inversions (2 and 4) and one translocation (1 and 5) have occurred after Vigna and Ceratotropis subgenera separation (~3.6 Mya). We also observed distinct oligopainting patterns for chromosomes 2 and 3 of Vigna species. Both Vigna species shared similar major rearrangements compared to Pv: one translocation (2 and 3) and one inversion (chromosome 3). The sequence synteny identified additional inversions and/or intrachromosomal translocations involving pericentromeric regions of both orthologous chromosomes. We propose chromosomes 2 and 3 as hotspots for chromosomal rearrangements and de novo centromere formation within and between Vigna and Phaseolus. Our BAC- and oligo-FISH mapping contributed to physically trace the chromosome evolution of Vigna and Phaseolus and its application in further studies of both genera.