Comparative Distribution of Repetitive Sequences in the Karyotypes of Xenopus tropicalis and Xenopus laevis (Anura, Pipidae)

Abstract

Xenopus laevis and its diploid relative, Xenopus tropicalis, are the most used amphibian models. Their genomes have been sequenced, and they are emerging as model organisms for research into disease mechanisms. Despite the growing knowledge on their genomes based on data obtained from massive genome sequencing, basic research on repetitive sequences in these species is lacking. This study conducted a comparative analysis of repetitive sequences in X. laevis and X. tropicalis. Genomic in situ hybridization (GISH) and fluorescence in situ hybridization (FISH) with Cot DNA of both species revealed a conserved enrichment of repetitive sequences at the ends of the chromosomes in these Xenopus species. The repeated sequences located on the short arm of chromosome 3 from X. tropicalis were not related to the sequences on the short arm of chromosomes 3L and 3S from X. laevis, although these chromosomes were homoeologous, indicating that these regions evolved independently in these species. Furthermore, all the other repetitive sequences in X. tropicalis and X. laevis may be species-specific, as they were not revealed in cross-species hybridizations. Painting experiments in X. laevis with chromosome 7 from X. tropicalis revealed shared sequences with the short arm of chromosome 3L. These regions could be related by the presence of the nucleolus organizer region (NOR) in both chromosomes, although the region revealed by chromosome painting in the short arm of chromosome 3L in X. laevis did not correspond to 18S + 28S rDNA sequences, as they did not colocalize. The identification of these repeated sequences is of interest as they provide an explanation to some problems already described in the genome assemblies of these species. Furthermore, the distribution of repetitive DNA in the genomes of X. laevis and X. tropicalis might be a valuable marker to assist us in understanding the genome evolution in a group characterized by numerous polyploidization events coupled with hybridizations.

Identification and characterization of a new family of long satellite DNA, specific of true toads (Anura, Amphibia, Bufonidae)

“…Chromosome preparations were obtained from bone marrow cells of adult individuals or from cell culture from tadpoles according to the protocols described in 75 .…”

Cytogenetic Analysis in the Toad Species Bufo spinosus, Bufotes viridis and Epidalea calamita (Anura, Bufonidae) from the Mediterranean Area

“…Tadpoles were euthanized by immersion in buffered 2% Tricaine methanesulfonate (MS 222; Sigma-Aldrich, Darmstadt, Germany) and then decapitated and dissected. Primary cell cultures were prepared from tadpole limbs as described in [ 28 ]. Secondary cell cultures were maintained in Dulbecco’s modified Eagle’s medium (DMEM) (Sigma-Aldrich, Taufkirchen, Germany) supplemented with 10% foetal bovine serum (PAA Laboratories, Cölbe, Germany), and 100 µg/mL penicillin, 100 U/mL streptomycin and 2.5 µg/mL amphotericin B (all antibiotics from Sigma-Aldrich, Darmstadt, Germany).…”

“…Probes were biotin-labelled for further use in FISH (rDNA and telomeric repeats) and GISH (genomic DNA) experiments. Telomeric repeats, (TTAGGG) n , were synthetized and labelled by PCR as described in [ 28 ]. One μg of recombinant plasmid (pDmr.a51) with a 11.5 kb insert encoding 18S and 28S ribosomal units from Drosophila melanogaster [ 32 ], was labelled using the Nick Translation Mix (Sigma-Aldrich, Darmstadt, Germany) according to the manufacturer’s instructions.…”

Divergent subgenome evolution in the allotetraploid frog Xenopus calcaratus

“…The allotetraploid species X. laevis has one NOR on chromosome 3L𝑝 (Tymowska and Kobel, 1972;Schmid et al, 1987;Roco, Liehr, Ruiz-García, Guzmán and Bullejos, 2021), with a homologous NOR on chromosome 3S thought to have been lost after allotetraploidization in subgenus Xenopus (Session et al, 2016). In the diploid X. tropicalis the NOR is on chromosome 7𝑞 (Tymowska, 1973;Tymowska and Fischberg, 1982;Uehara, Haramoto, Sekizaki, Takahashi and Asashima, 2002;Roco et al, 2021), the allotetraploid X. epitropicalis and X. mellotropicalis also have one NOR on chromosome 7a𝑞 (Tymowska and Fischberg, 1982;Tymowska, 1991), with the other on chromosome 7b hypothesized to have been lost after allotetraploidization (Knytl et al, 2017). In X. calcaratus, 28S rDNA FISH identified nucleolar (secondary) constriction on XCA 7a𝑞 (Figs.…”