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Molecular cloning and characterization of satellite DNA sequences from constitutive heterochromatin of the habu snake (Protobothrops flavoviridis, Viperidae) and the Burmese python (Python bivittatus, Pythonidae)

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Abstract

Highly repetitive DNA sequences of the centromeric heterochromatin provide valuable molecular cytogenetic markers for the investigation of genomic compartmentalization in the macrochromosomes and microchromosomes of sauropsids. Here, the relationship between centromeric heterochromatin and karyotype evolution was examined using cloned repetitive DNA sequences from two snake species, the habu snake (Protobothrops flavoviridis, Crotalinae, Viperidae) and Burmese python (Python bivittatus, Pythonidae). Three satellite DNA (stDNA) families were isolated from the heterochromatin of these snakes: 168-bp PFL-MspI from P. flavoviridis and 196-bp PBI-DdeI and 174-bp PBI-MspI from P. bivittatus. The PFL-MspI and PBI-DdeI sequences were localized to the centromeric regions of most chromosomes in the respective species, suggesting that the two sequences were the major components of the centromeric heterochromatin in these organisms. The PBI-MspI sequence was localized to the pericentromeric region of four chromosome pairs. The PFL-MspI and the PBI-DdeI sequences were conserved only in the genome of closely related species, Gloydius blomhoffii (Crotalinae) and Python molurus, respectively, although their locations on the chromosomes were slightly different. In contrast, the PBI-MspI sequence was also in the genomes of P. molurus and Boa constrictor (Boidae), and additionally localized to the centromeric regions of eight chromosome pairs in B. constrictor, suggesting that this sequence originated in the genome of a common ancestor of Pythonidae and Boidae, approximately 86 million years ago. The three stDNA sequences showed no genomic compartmentalization between the macrochromosomes and microchromosomes, suggesting that homogenization of the centromeric and/or pericentromeric stDNA sequences occurred in the macrochromosomes and microchromosomes of these snakes.

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Acknowledgments

We are grateful to Michihisa Toriba, Japan Snake Institute, Gunma, and Yoshinori Kumazawa, Nagoya City University, for kindly providing snake samples.

Funding

This research was partially supported by Grants-in-Aid for Scientific Research on Innovative Areas (no. 23113004) and a Grant-in-Aid for Scientific Research (B) (no. 22370081) from the Ministry of Education, Culture, Sports, Science and Technology, Japan.

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

Animal care and all experimental procedures were approved by the Animal Experiment Committee (Approval no. CAST05-003), Graduate School of Science, Hokkaido University, and the experiments were conducted according to the Regulations on Animal Experiments in Hokkaido University.

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Authors and Affiliations

  1. Laboratory of Animal Genetics, Department of Applied Molecular Biosciences, Graduate School of Bioagricultural Sciences, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, Aichi, 464-8601, Japan

    Kazumi Matsubara, Yoshinobu Uno, Kornsorn Srikulnath & Yoichi Matsuda

  2. Laboratory of Animal Cytogenetics and Comparative Genomics, Department of Genetics, Faculty of Science, Kasetsart University, 50 Ngamwongwan, Chatuchak, Bangkok, 10900, Thailand

    Kornsorn Srikulnath

  3. Center for Advanced Studies in Tropical Natural Resources (CASTNAR), National Research University-Kasetsart University (NRU-KU), Kasetsart University, Bangkok, Thailand

    Kornsorn Srikulnath

  4. Laboratory of Animal Cytogenetics, Biosystems Science Course, Graduate School of Life Science, Hokkaido University, North 10 West 8, Kita-ku, Sapporo, 060-0810, Japan

    Risako Seki

  5. Department of Biological Sciences, Faculty of Science, Hokkaido University, North 10 West 8, Kita-ku, Sapporo, 060-0810, Japan

    Chizuko Nishida

  6. Avian Bioscience Research Center, Graduate School of Bioagricultural Sciences, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, 464-8601, Japan

    Yoichi Matsuda

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  1. Kazumi Matsubara
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  2. Yoshinobu Uno
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Correspondence to Kazumi Matsubara.

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Supplementary Figure 1

Nucleotide sequences of repeat sequence families. Alignments of 22 PFL-MspI fragments (A), 32 PBI-DdeI fragments (B) and 3 PBI-MspI fragments (C). Dots indicate the same nucleotides as those of the consensus sequence of each sequence family, and the hyphens indicate gaps. Internal restriction sites of eight endonucleases are indicated as follows: (PDF 73 kb)

AluI (GIF 812 b)

ApaI (GIF 842 b)

DdeI (GIF 883 b)

EcoT22I (GIF 929 b)

HaeIII (GIF 829 b)

HinfI (GIF 818 b)

MspI (HpaII) (GIF 827 b) and

Sau3AI (GIF 842 b).

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Matsubara, K., Uno, Y., Srikulnath, K. et al. Molecular cloning and characterization of satellite DNA sequences from constitutive heterochromatin of the habu snake (Protobothrops flavoviridis, Viperidae) and the Burmese python (Python bivittatus, Pythonidae). Chromosoma 124, 529–539 (2015). https://doi.org/10.1007/s00412-015-0529-6

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