Isolation and characterization of 5S rDNA sequences in catfishes genome (Heptapteridae and Pseudopimelodidae): perspectives for rDNA studies in fish by C0t method

Abstract

Sequences of 5S ribosomal RNA (rRNA) are extensively used in fish cytogenomic studies, once they have a flexible organization at the chromosomal level, showing inter- and intra-specific variation in number and position in karyotypes. Sequences from the genome of Imparfinis schubarti (Heptapteridae) were isolated, aiming to understand the organization of 5S rDNA families in the fish genome. The isolation of 5S rDNA from the genome of I. schubarti was carried out by reassociation kinetics (C0t) and PCR amplification. The obtained sequences were cloned for the construction of a micro-library. The obtained clones were sequenced and hybridized in I. schubarti and Microglanis cottoides (Pseudopimelodidae) for chromosome mapping. An analysis of the sequence alignments with other fish groups was accomplished. Both methods were effective when using 5S rDNA for hybridization in I. schubarti genome. However, the C0t method enabled the use of a complete 5S rRNA gene, which was also successful in the hybridization of M. cottoides. Nevertheless, this gene was obtained only partially by PCR. The hybridization results and sequence analyses showed that intact 5S regions are more appropriate for the probe operation, due to conserved structure and motifs. This study contributes to a better understanding of the organization of multigene families in catfish’s genomes.

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Acknowledgments

The authors are grateful to Dr. André Luis LaforgaVanzela and Dra. Maria Helena Pelegrinelli Fungaro for support to cloning in this study. This research was supported by a grant from Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES). This search received permission from Instituto Brasileiro do Meio Ambiente e dos Recursos Naturais Renováveis (IBAMA) to collect fish specimens.

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Correspondence to Ana Lúcia Dias.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material S1

PCR amplification of 5S rDNA of Imparfinis schubarti genome in 1 % agarose gel (PDF 95 kb)

Supplementary material S2

Results of CENSOR software shows in sequences with incomplete 5S, similar to the SINE3-1 element (PDF 46 kb)

Supplementary material S3

Prediction of the secondary structure of the 5S rRNA obtained by the C0t method. The color scale indicates the likelihood of the pairings (PDF 106 kb)

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Gouveia, J.G., Wolf, I.R., de Moraes-Manécolo, V.P.O. et al. Isolation and characterization of 5S rDNA sequences in catfishes genome (Heptapteridae and Pseudopimelodidae): perspectives for rDNA studies in fish by C0t method. Cytotechnology 68, 2711–2720 (2016). https://doi.org/10.1007/s10616-016-9996-8

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Keywords

  • C0t
  • Genomics micro-library
  • Hybridization
  • Pisces