Karyotype Differentiation in Cultivated Chickpea Revealed by Oligopainting Fluorescence in situ Hybridization

Abstract

Chickpea (Cicer arietinum L.) is one of the main sources of plant proteins in the Indian subcontinent and West Asia, where two different morphotypes, desi and kabuli, are grown. Despite the progress in genome mapping and sequencing, the knowledge of the chickpea genome at the chromosomal level, including the long-range molecular chromosome organization, is limited. Earlier cytogenetic studies in chickpea suffered from a limited number of cytogenetic landmarks and did not permit to identify individual chromosomes in the metaphase spreads or to anchor pseudomolecules to chromosomes in situ. In this study, we developed a system for fast molecular karyotyping for both morphotypes of cultivated chickpea. We demonstrate that even draft genome sequences are adequate to develop oligo-fluorescence in situ hybridization (FISH) barcodes for the identification of chromosomes and comparative analysis among closely related chickpea genotypes. Our results show the potential of oligo-FISH barcoding for the identification of structural changes in chromosomes, which accompanied genome diversification among chickpea cultivars. Moreover, oligo-FISH barcoding in chickpea pointed out some problematic, most probably wrongly assembled regions of the pseudomolecules of both kabuli and desi reference genomes. Thus, oligo-FISH appears as a powerful tool not only for comparative karyotyping but also for the validation of genome assemblies.

Oligo-barcode illuminates holocentric karyotype evolution in Rhynchospora (Cyperaceae)

“…However, most plant groups still have only a single reference genome, making comparative genomic analyses between closely related species difficult and requiring the use of alternative feasible methods ( Parween et al., 2015 ; Qin et al., 2019 ; Escudero et al., 2023 ). Chromosome barcoding with oligo-based probes has been used to determine karyotypes and investigate chromosomal rearrangements, meiotic pairing, and recombination in a wide range of species ( Braz et al., 2018 ; Meng et al., 2018 ; de Oliveira Bustamante et al., 2021 ; do Vale Martins et al., 2021 ; Li et al., 2021 ; Doležalová et al., 2022 ; Nascimento and Pedrosa-Harand, 2023 ). Oligo-FISH barcoding is based on the production of two different oligomer libraries from multiple regions of multiple chromosomes to produce an unique barcode signal pattern for each individual chromosome pair, facilitating the unambiguous identification of all chromosomes of a species in a single FISH experiment ( Braz et al., 2018 ; Braz et al., 2020 ; de Oliveira Bustamante et al., 2021 ; Liu and Zhang, 2021 ).…”

Tracing the Evolution of the Angiosperm Genome from the Cytogenetic Point of View

“…An oligo pool that is designed from a single-copy DNA sequence can be used as hybridisation probes that are specific to a chromosomal region (or regions) [ 87 ], to a whole chromosome arm [ 102 ] or to an entire chromosome [ 103 ]. Recently, oligo–FISH barcoding pools were developed for several mainly cultivated species, which enabled the detection of various chromosomal rearrangements that accompanied their evolution and speciation [ 87 , 104 , 105 ]. However, there are some taxa such as Citrus and Populus that seem to have very conserved karyograms.…”

“…However, in others the interstitial telomeric sequences could be inserted via a translocation with mobile elements or by a mechanism of the rolling-circle replication of extrachromosomal circular DNA [ 62 , 107 , 114 ]. Regardless of its origin, when the interstitial locus of telomeric repeats exists, it is often a very good chromosome marker [ 104 ]. The rDNA sequences and telomeric repeats are evolutionary conserved and, once they are isolated and cloned, can be used to analyse a wide variety of plant species while satellite repeats are specific to a species or to a small groups of closely related taxa [ 107 , 115 ].…”

Oligo-barcodes illuminate holocentric karyotype evolution inRhynchospora(Cyperaceae)

“…However, most plant groups still have only a single reference genome, making comparative genomic analyses between closely related species difficult and requiring the use of alternative feasible methods (Parween et al, 2015;Qin et al, 2019;Escudero et al, 2023). Chromosome barcoding with oligo-based probes has been used to determine karyotypes and investigate chromosomal rearrangements, meiotic pairing, and recombination in a wide range of species (Braz et al, 2018;Meng et al, 2018;Li et al, 2021;Do Vale Martins et al, 2021;De Oliveira Bustamante et al, 2021;Doležalová et al, 2022;Nascimento and Pedrosa-Harand, 2023). Oligo-FISH barcoding is based on the production of two different oligomer libraries from multiple regions of multiple chromosomes to produce an unique barcode signal pattern for each individual chromosome pair, facilitating the unambiguous identification of all chromosomes of a species in a single FISH experiment (Braz et al, 2018(Braz et al, , 2020Liu and Zhang 2021;De Oliveira Bustamante et al, 2021).…”