bioRxiv preprint first posted online Mar. 24, 2016; doi: http://dx.doi.org/10.1101/042945.

The copyright holder for this preprint (which was

not peer-reviewed) is the author/funder. It is made available under a CC-BY-ND 4.0 International license.

bioRiv Poliseno et al. 2016. bioRiv

New Results
Complete mitochondrial genome of Muricea crassa and
Muricea purpurea (Anthozoa: Octocorallia) from the
eastern tropical Pacific
Angelo Poliseno1 , Odalisca Breedy2,3,4 , Michael Eitel1 , Gert
Wrheide1,5,6 , Hector M. Guzman4 , Stefan Krebs7 , Helmuth Blum7 , and
Sergio Vargas1
1 Department of Earth and Environmental Sciences, Palaeontology & Geobiology,
Ludwig-Maximilians-Universitt Mnchen, Richard-Wagner-Strae 10, 80333 Mnchen, Germany
2 Centro de Investigacin en Estructuras Microscpicas, Universidad de Costa Rica. P.O. Box 11501-2060,

Universidad de Costa Rica, San Jos, Costa Rica

3 Centro de Investigaci
n en Ciencias del Mar y Limnologa, Escuela de Biologa, Universidad de Costa Rica.
P.O. Box 11501-2060, Universidad de Costa Rica, San Jos, Costa Rica
4 Smithsonian Tropical Research Institute, P.O. Box 0843-03092 Panama, Panama
5 GeoBio-Center, Ludwig-Maximilians-Universitt Mnchen, Richard-Wagner-Strae 10, 80333 Mnchen,

Germany
6 Bayerische Staatssammlung fr Palontologie und Geologie, Richard-Wagner-Strae 10, 80333 Mnchen,

Germany
7 Laboratory for Functional Genome Analysis (LAFUGA), Gene Center, Ludwig-Maximilians-Universitt

Mnchen, Feodor-Lynen-Strasse 25, 81377 Mnchen, Germany

Abstract
We sequenced the complete mitogenomes of two eastern tropical Pacific gorgonians,
Muricea crassa and Muricea purpurea, using NGS technologies. The assembled mi-
togenomes of M. crassa and M. purpurea were 19,586 bp and 19,358 bp in length,
with a GC-content ranging from 36.0% to 36.1%, respectively. The two mitogenomes
had the same gene arrangement consisting of 14 protein-coding genes, two rRNAs and
one tRNA. Mitogenome identity was 98.5%. The intergenic regions between COB and
NAD6 and between NAD5 and NAD4 were polymorphic in length with a high level of nu-
cleotide diversity. Based on a concatenated dataset of 14 mitochondrial protein-coding
genes we inferred the phylogeny of 26 octocoral species.

Key words: mitogenome, Muricea crassa, Muricea purpurea, NGS

Muricea crassa (Verrill, 1869) and Muricea purpurea (Verrill, 1864) are two
shallow water gorgonians of the family Plexauridae. Their distribution is limited
to the eastern tropical Pacific where they are abundant members of coral com-
munities and littoral zones (Guzman et al., 2004). Samples were collected as
part of an ecological and biodiversity survey undertaken in the Coiba National
Park (Panama). Genomic DNA was extracted from ethanol-preserved samples
Preprint

and was used to construct genomic libraries using the Accel-NGS 1S DNA Li-
brary kit (Swift Biosciences, Ann Arbor, MI, USA) following the manufacturers
instructions. These libraries were sequenced (100bp PE) on an Illumina HiSeq

Corresponding author: sergio.vargas@lmu.de

cbd
bioRxiv preprint first posted online Mar. 24, 2016; doi: http://dx.doi.org/10.1101/042945. The copyright holder for this preprint (which was
not peer-reviewed) is the author/funder. It is made available under a CC-BY-ND 4.0 International license.

Poliseno et al. 2016 Eastern Pacific Muricea Mitogenomes

(Illumina Inc., San Diego, CA). The quality of the reads obtained was assessed
with FastQC (Andrews, 2010), low quality reads and Illumina adaptors were
trimmed using Trimmomatic 0.3.2 called from Trinity RNA-Seq 2.0.6 (Grabherr
et al., 2011). Despite its original purpose, the Trinity RNA-Seq assembler was
used after normalization to 50X coverage for de-novo mitogenome assembly. The
assembly resulted in a single mitochondrial contig in both species. Initial anno-
tation was performed with the ORF finder function implemented in Geneious
8.1.7 (Kearse et al., 2012) and was corroborated by comparison with published
octocoral mitogenomes. The presence of DNA repeats was assessed with the tan-
dem repeats finder server 4.08 available at https://tandem.bu.edu/trf/trf.html
(Benson, 1999). The complete mitogenomes of M. crassa (LT174652) and M.
purpurea (LT174653) were 19,586 bp and 19,358 bp long, with a GC-content
of 36.0% (M. purpurea) and 36.1% (M. crassa), respectively. Both mitogenomes
had gene arrangement of type A (see Brockman and McFadden, 2012). In to-
tal, the Coding DNA Sequences (CDSs) spanned about 76% of the mitogenome
in both species. Among protein-coding genes, the highest level of nucleotide
diversity (0.4%) was found in NAD1, NAD6 and COX2, whereas no nucleotide
substitutions were found in NAD3, ATP6 and ATP8. Except for NAD2 and NAD5
(13bp overlap), the other protein-coding genes were separated by intergenic re-
gions (IGRs) of different lengths. In both species, the shortest IGRs were those
located between 12S rRNA and NAD1 and between 16S rRNA and NAD2, while
the longest was found between NAD5 and NAD4. The latter IGR was also the
most diverse region with a nucleotide diversity of 6.2%. Length polymorphism
was found in the COB-NAD6 IGR, which was 184bp shorter in M. purpurea than
in M. crassa. Sequencing of indel-rich IGRs such as that between COB and NAD6
may result useful for molecular species-identification in the genus Muricea. Fi-
nally, we found a 37 bp tandem repeat in the IGR between NAD4 and tRNA of
M. crassa.
The two newly sequenced complete mitogenomes were used to assess the
phylogenetic relationships among 26 different octocoral species. A concatenated
nucleotide alignment of 14 protein-coding genes (15,249 bp in total) for 41 taxa
was generated with MUSCLE (Edgar, 2004) using the default options provided
in Seaview (Gouy et al., 2010). The maximum likelihood tree was inferred in
RAxML 7.2.8 (Stamatakis, 2006) under the GTR+GAMMA substitution model.
Node support was estimated using 1000 bootstrap pseudoreplicates. The phy-
logenetic tree (Figure 1) was re-rooted using three calcaxonians and two pen-
Preprint

natulaceans as outgroup (not shown in Figure 1). Tree topology is consistent

with recently published studies (Figueroa and Baco, 2015). The phylogenetic
placement of Muricea spp. sister to Nephtheidae (Dendronephthya spp. and Scle-
ronephthya spp.) is likely an artifact caused by poor taxon sampling in the family
Plexauridae.
cbd 2 bioRiv
bioRxiv preprint first posted online Mar. 24, 2016; doi: http://dx.doi.org/10.1101/042945. The copyright holder for this preprint (which was
not peer-reviewed) is the author/funder. It is made available under a CC-BY-ND 4.0 International license.

Poliseno et al. 2016 Eastern Pacific Muricea Mitogenomes

Acknowledgements
We would like to thank the Gene Center (Ludwig-Maximilians-Universitt, Mnchen)
for technical support and sequencing. SV is indebted to N. Villalobos, M. Vargas
and S. Vargas for their constant support.

Declaration of interest
The authors report no conflicts of interest. The authors alone are responsible for
the content and writing of the paper. The project was partially supported by the
LMU Mnchen German Excellence Initiative Junior Research Funds to SV, the
Smithsonian Tropical Research Institute and the Secretara Nacional de Ciencias
y Tecnologa (SENACYT) de Panam to HMG. Collection and exporting permits
of eastern Pacific material were issued by the Autoridad de Recursos Marinos de
Panam and by the Ministerio de Ambiente de Panam.

References
Andrews S. (2010). FastQC: a quality control tool for high throughput se-
quence data. Available online at: http://www.bioinformatics.babraham.ac.
uk/projects/fastqc

Benson G. (1999). Tandem repeats finder: a program to analyze DNA sequences.

Nucleic Acids Res 27:573-580.

Brockman SA, McFadden CS. (2012). The Mitochondrial Genome of Param-

inabea aldersladei (Cnidaria: Anthozoa: Octocorallia) supports intramolecular
recombination as the primary mechanism of gene rearrangement in octocoral
mitochondrial genomes. Genome Biol Evol 4:994-1006.

Edgar RC. (2004). MUSCLE: multiple sequence alignment with high accuracy
and high throughput. Nucleic Acids Res 32:1792-1797.

Figueroa FF, Baco AR. (2015). Octocoral mitochondrial genomes provide in-
sights into the phylogenetic history of gene order rearrangements, order rever-
Preprint

sals, and cnidarian phylogenetics. Genome Biol Evol 7:391-409.

cbd 3 bioRiv
bioRxiv preprint first posted online Mar. 24, 2016; doi: http://dx.doi.org/10.1101/042945. The copyright holder for this preprint (which was
not peer-reviewed) is the author/funder. It is made available under a CC-BY-ND 4.0 International license.

Poliseno et al. 2016 Eastern Pacific Muricea Mitogenomes

Gouy M, Guindon S, Gascuel O. (2010). SeaView version 4: a multiplatform

graphical user interface for sequence alignment and phylogenetic tree building.
Mol Biol Evol 27:221-224.

Grabherr MG, Haas BJ, Yassour M, Levin JZ, Thompson DA, Amit I, Adiconis X,
et al. (2011). Full-length transcriptome assembly from RNA-Seq data without a
reference genome. Nature Biotechnol 29:644-652.

Guzman HM, Guevara CA, Breedy O. (2004). Distribution, diversity, and conser-
vation of coral reefs and coral communities in the largest marine protected area
of Pacific Panama (Coiba Island). Environ Conserv 31:111-121.

Kearse M, Moir R, Wilson A, Stones-Havas S, Cheung M, Sturrock S, Buxton S,

et al. (2012). Geneious Basic: an integrated and extendable desktop software
platform for the organization and analysis of sequence data. Bioinformatics 28:
1647-1649.

Stamatakis A. (2006). RAxML-VI-HPC: maximum likelihood based phylogenetic

analyses with thousands of taxa and mixed models. Bioinformatics 22:2688-
2690.

Preprint

cbd 4 bioRiv
 cbd
JX508792 Paraminabea aldersladei
100 KM015352 Anthomastus sp. .
KM015353 Anthomastus sp.
100 KM015354 Sibogagorgia cauliflora
100 AB595189 Paracorallium japonicum
100 100 AB700136 Corallium rubrum
Poliseno et al. 2016

100 100 KC782352 Corallium imperiale

100 KC782348 Corallium laauense
KC782355 Corallium imperiale
KC782353 Corallium kishinouyei
100
100 KC782347 Corallium secundum
100
100 AB595190 Corallium konojoi
AB700134 Corallium elatius
100 AB700135 Corallium elatius
100 KC782351 Paragorgia sp.
100 KC782356 Paragorgia sp.
100 KC782350 Paragorgia sp.
98 KC782354 Paragorgia sp.
100 KC782349 Paragorgia sp.
KF785800 Paragorgia coralloides
93 KF785801 Paragorgia coralloides
DQ640649 Briareum asbestinum
100 JX023274 Sinularia peculiaris
78 HQ694728 Euplexaura crassa
DQ640646 Pseudopterogorgia bipinnata

5
100
GU047879 Scleronephthya gracillimum
100 100 JQ290079 Dendronephthya suensoni
100 100 HQ694726 Dendronephthya putteri
GU047878 Dendronephthya suensoni
100 FJ372991 Dendronephthya gigantea
97 HQ694725 Dendronephthya mollis
100 GU047877 Dendronephthya castanea
72 HQ694727 Echinogorgia complexa
100 LT174653 Muricea purpurea
LT174652 Muricea crassa

0.02

Figure 1: Phylogenetic tree of 41 octocorals based on a concatenated alignment of 14 mitochondrial protein-coding genes. The calcaxonians
(Keratoisidinae sp., Acanella eburnea, Narella hawaiinensis and Junceella fragilis) and pennatulaceans (Renilla muelleri and Stylatula elongata)
were used to re-root the tree but are not shown here. Numbers at the nodes indicate bootstrap values.
not peer-reviewed) is the author/funder. It is made available under a CC-BY-ND 4.0 International license.

bioRiv
Eastern Pacific Muricea Mitogenomes
bioRxiv preprint first posted online Mar. 24, 2016; doi: http://dx.doi.org/10.1101/042945. The copyright holder for this preprint (which was

Preprint