Elsevier

Gene

Volume 713, 10 September 2019, 143967
Gene

Research paper
Diagnosis of mitogenome for robust phylogeny: A case of Cypriniformes fish group

https://doi.org/10.1016/j.gene.2019.143967 Get rights and content

Highlights

  • We report the first complete mitogenome of Queen loach, Botia dario using next-generation sequencing methods.

  • Phylogenetics analysis of complete mitogenome of B. dario and some related fish species from the order Cypriniformes.

  • A study on Cypriniformes based on complete mitogenome and individual mitogenes to find a suitable gene for phylogenetic tree construction.

Abstract

Phylogenetic tree using mitochondrial genes and nuclear genes have long been used for augmenting biological classification and understanding evolutionary processes in different lineage of life. But a basic question still exists for finding the most suitable gene for constructing robust phylogenetic tree. Much of the controversy appears due to monophyletic, paraphyletic and polyphyletic clade making deviations from original taxonomy. In the present study we report the first complete mitochondrial genome (mitogenome) of queen loach, generated through next-generation sequencing methods. The assembled mitogenome is a 16,492 bp circular DNA, comprising of 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes and a control region. Further in this study we also investigated the suitability of different mitochondrial region for phylogenetic analysis in Cyprinidae and loach group. For this genetic tree were constructed on COI, COII, COIII, 16S rRNA, 12S rRNA, Cyt b, ATPase 6, D-loop, ND1, ND2, ND3, ND4, ND5, and ND6 along with complete mitogenome. The complete mitogenome based phylogenetic tree got inclusive support from available classical taxonomy for these groups. On individual gene basis Cyt b, 12S rRNA, ND2 and ND3 also produced perfect clade at family and subfamily level. For rest of the genes polyphyly were observed for the fishes belonging to same family or subfamily which makes their use questionable for phylogenetic tree construction.

Introduction

The phylogeny based on molecular data are being used extensively for understanding and placing more accurately the evolution of gene and genome in conjunction with origin and divergence of species (Brown, 2002). But not limited to this scope, phylogenetics is also being used in the field of understanding molecular mechanism of disease (Harris et al., 2010), development of medicine (Espiritu et al., 2001; Fenical et al., 2009), Bioprospecting (Smith and Wheeler, 2006), organisms' metabolic networks (Forst and Schulten, 2001), etc. However, different genes such as mitochondrial (e.g. COI, Cyt-b, 16S rRNA) and nuclear (e.g.RAG, ATPS-α) are used for studying evolutionary history, produces different kind of topology or clade in phylogenetic tree for different group of life and further inference (Castresana, 2007; Powell et al., 2013; Farias et al., 2001). Since the process or technique of drawing phylogenetic relationship among the species is still establishing, hunt for a better gene continues. However, this is not so straightforward, as we analyses the final established sequence for different regions (genes) which has already gone through some convergent, divergent and independent evolution in different branches of life for better adaptation, selection and speciation (Zuckerkandl and Pauling, 1965; Zakon, 2002; Hasselmann et al., 2008; Castoe et al., 2009; Arbuckle and Speed, 2016) resulting in contrasting phylogenetic tree. Hence, in the present study, we compared and evaluated different mitochondrial regions along with complete mitogenome for making the phylogenetic tree in some selected group of fishes with concurrence note with the classical taxonomic position. For this purpose, we also sequenced complete mitochondrial DNA of the queen loach, Botia dario.

B. dario belongs to the family Botiidae of order Cypriniformes and found in Brahmaputra and Ganges basins of Bangladesh, Bhutan and Northeast India (Talwar and Jhingran, 1991). The species has a great demand for the aquarium trade in many countries due to its unique band pattern and bright color (Froese and Pauly, 2015). This species is considered to be endangered in Bangladesh (IUCN Bangladesh, 2000) and near endangered in lower Brahmaputra basin of Assam, India due to habitat destruction and overexploitation for ornamental trade (CAMP, 1998). So its conservation measures are of a great concern now.

Section snippets

Ethical statement

All the fish specimens of B. dario used in this study were collected from the College of Fisheries, CAU (I) tank. Ethical approval, specimen collection, and maintenance were performed in strict accordance with the recommendations of the ethical standards of the Institute of Animal Ethics Committee, College of Fisheries, Central Agricultural University, Imphal.

Tissue collection and DNA isolation

The mitochondrial DNA, used in this study, was isolated from muscle tissue of B. dario fish species, after following the method of

Characteristics of the B. dario mitogenome

We determined the first complete mitochondrial genome sequence of B. dario by next-generation sequencing method. A uniform coverage of ~1500× was obtained for most of the mitogenome segments, except a few places in the control region where the presence of the repeat sequences resulted in slightly low coverage. A total of 31,52,420 reads were found from the mitochondrial DNA sample, used with a total data size of approximately 939 Mb. Size of the assembled mitogenome, as generated from CLC

Discussion

The fishes of Cobitidae, Nemacheilidae and Balitoridae family clearly formed a separate clade in the phylogenetic tree based on complete mitogenome (Fig. 3). All the three families have very similar morphometric characters and commonly known as loach. The other species of Cyprinidae family formed a paraphyletic clade with loach group which exhibits the evolutionary relatedness and sharing of a common ancestor between Cyprinidae and loach group. This finding is in agreement with a previous

Conclusion

In the present study, the complete mitochondrial DNA sequence of B. dario found will help in studying its biology, molecular studies, evolutionary studies, species confirmation and will be useful in conservation management. The whole mitogenome will also help in population studies of this fish species. The complete mitogenome of B. dario found in this study has been submitted to NCBI GenBank with accession number MG751071.The phylogenetic analysis of individual mitochondrial genes along with

Declaration of Competing Interest

The authors declare no conflict of interest.

Acknowledgments

The authors thank the Centre of Excellence (COE) (Grant number- DBT-NER/LIVE/05/2011) and Department of Biotechnology, Government of India (Grant number- BT/22/NE/2011) for providing instruments for the experiment.

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