Draft genome of the Northern snakehead, Channa argus

TLDR: A high-quality draft genome of C. argus is generated, which will provide a valuable genetic resource for further biomedical investigations of this economically important teleost fish.

Abstract: Background: The Northern snakehead (Channa argus), a member of the Channidae family of the Perciformes, is an economically important freshwater fish native to East Asia. In North America, it has become notorious as an intentionally released invasive species. Its ability to breathe air with gills and migrate short distances over land makes it a good model for bimodal breath research. Therefore, recent research has focused on the identification of relevant candidate genes. Here, we performed whole genome sequencing of C. argus to construct its draft genome, aiming to offer useful information for further functional studies and identification of target genes related to its unusual facultative air breathing. Findings: We assembled the C. argus genome with a total of 140.3 Gb of raw reads, which were sequenced using the Illumina HiSeq2000 platform. The final draft genome assembly was approximately 615.3 Mb, with a contig N50 of 81.4 kb and scaffold N50 of 4.5 Mb. The identified repeat sequences account for 18.9% of the whole genome. The 19 877 protein-coding genes were predicted from the genome assembly, with an average of 10.5 exons per gene. Conclusion: We generated a high-quality draft genome of C. argus, which will provide a valuable genetic resource for further biomedical investigations of this economically important teleost fish.

Full text

Giga Science, 6, 2017, 1–5
doi: 10.1093/gigascience/gix011
Advance Access Publication Date: 2 March 2017
Data Note
DATA NOTE
Draft genome of the Northern snakehead, Channa
argus
Jian Xu
1,†
, Chao Bian
2,3,4,†
, Kunci Chen
5,†
, Guiming Liu
6
, Yanliang Jiang
1
,
Qing Luo
5
, Xinxin You
2,3
, Wenzhu Peng
1,7
,JiaLi
3
,YuHuang
3
, Yunhai Yi
3
,
Chuanju Dong
1,8
,HuaDeng
9
, Songhao Zhang
1
, Hanyuan Zhang
1
,
Qiong Shi
2,3,10,∗
and Peng Xu
1,7,∗
1
Key Laboratory of Aquatic Genomics, Ministry of Agriculture, CAFS Key Laboratory of Aquatic Genomics and
Beijing Key Laboratory of Fishery Biotechnology, Chinese Academy of Fishery Sciences, Fengtai, Beijing,
100141, China,
2
BGI Research Center for Aquatic Genomics, Chinese Academy of Fishery Sciences, Shenzhen,
Guangdong, 518083, China,
3
Shenzhen Key Lab of Marine Genomics, Guangdong Provincial Key Lab of
Molecular Breeding in Marine Economic Animals, BGI, Shenzhen, Guangdong, 518083, China,
4
Centre of
Reproduction, Development and Aging, Faculty of Health Sciences, University of Macau, Taipa, Macau, China,
5
Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou, Guangdong,
510380, China,
6
CAS Key Laboratory of Genome Sciences and Information, Beijing Institute of Genomics,
Chinese Academy of Sciences, Chaoyang, Beijing, 100029, China,
7
Fujian Collaborative Innovation Center for
Exploitation and Utilization of Marine Biological Resources, Xiamen University, Xiamen, Fujian, 361102, China,
8
College of Fishery, Henan Normal University, Xinxiang, Henan, 453007, China,
9
Research Institute of Forestry
Policy and Information,Chinese Academy of Forestry, Haidian, Beijing, 100091, China and
10
Laboratory of
Aquatic Genomics, College of Ecology and Evolution, School of Life Sciences, Sun Yat-Sen University,
Guangzhou, Guangdong, 510275, China
∗
Correspondence address. Qiong Shi: BGI Research Center for Aquatic Genomics, Chinese Academy of Fishery Sciences, Shenzhen, Guangdong, 518083,
China. E-mail: shiqiong@genomics.cn; Peng Xu: Key Laboratory of Aquatic Genomics, Ministry of Agriculture, CAFS Key Laboratory of Aquatic Genomics
and Beijing Key Laboratory of Fishery Biotechnology, Chinese Academy of Fishery Sciences, Fengtai, Beijing, 100141, China.
E-mail: xupeng77@xmu.edu.cn
†
Contributed equally to this work.
Abstract
Background: The Northern snakehead (Channa argus), a member of the Channidae family of the Perciformes, is an
economically important freshwater sh native to East Asia. In North America, it has become notorious as an intentionally
released invasive species. Its ability to breathe air with gills and migrate short distances over land makes it a good model
for bimodal breath research. Therefore, recent research has focused on the identication of relevant candidate genes. Here,
we performed whole genome sequencing of C. argus to construct its draft genome, aiming to offer useful information for
Received: 17 August 2016; Revised: 5 January 2017; Accepted: 25 February 2017
C

The Author 2017. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons
Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium,
provided the original work is properly cited.
1
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2 Xu et al.
further functional studies and identication of target genes related to its unusual facultative air breathing. Findings: We
assembled the C. argus genome with a total of 140.3 Gb of raw reads, which were sequenced using the Illumina HiSeq2000
platform. The nal draft genome assembly was approximately 615.3 Mb, with a contig N50 of 81.4 kb and scaffold N50 of 4.5
Mb. The identied repeat sequences account for 18.9% of the whole genome. The 19 877 protein-coding genes were
predicted from the genome assembly, with an average of 10.5 exons per gene. Conclusion: We generated a high-quality
draft genome of C. argus, which will provide a valuable genetic resource for further biomedical investigations of this
economically important teleost sh.
Keywords: Channa argus; genome assembly; annotation; gene prediction
Data description
Introduction of C. argus
The Northern snakehead (Channa argus) is a special snakehead
sh cultivated mainly in Asia and Africa for food, especially in
China with an annual production of about 510 000 tons (worth
∼1.6 billion US dollars) (Fig. 1). Genetic degradation caused by in-
breeding of C. argus cultivation has led to higher susceptibility to
diseases. Furthermore, C. argus is considered a serious invasive
species in North America, due to its wide-range diet, parental
care, and rapid colonization and expansion [1]. C. argus has a
specialized aerial breathing organ, the suprabranchial chamber,
which facilitates its aquatic–aerial bimodal breathing. Because
of its aggressive status in ecosystem of rivers, lakes, and ponds,
and little consumption of the C. argus in America for food, this
leads to threats to the balance of ecosystems. For both economic
and ecological consideration, it is vital to develop genomic
resources for further genetic breeding studies or ecological re-
search. So far, the genome sequence of C. argus has not been
reported, and hence in our current study we performed genome
sequencing, assembly, and annotation of this teleost species.
C. argus genome sequencing on the Illumina platform
Genomic DNA was extracted from blood sample of a single fe-
male C. argus (Fishbase ID: 4799) using Qiagen GenomicTip100
(Qiagen). The sh was obtained from the Pearl River Fish-
eries Research Institute, Chinese Academy of Fishery Sciences,
Guangzhou, China. A whole-genome shotgun sequencing strat-
egy was applied, and short-insert libraries (180, 500, and 800
bp) and long-insert libraries (3 and 5 kb) were constructed us-
ing the standard protocol provided by Illumina (San Diego, CA,
USA). Paired-end sequencing with a 2 × 100-bp read length was
Figure: 1: the Northern snakehead sh, Channa argus.
performed on the short-insert and long-insert libraries using the
Illumina HiSeq2000 platform. In total, we generated about 140.3
Gb of raw reads, including 33.0, 36.9, 17.4, 26.5, and 26.5 Gb of
reads from the 180-, 500-, 800-, 3-, and 5-kb libraries. After re-
moval of low-quality and redundant reads, we obtained about
138.2 Gb of clean data for further de novo assembling of the C.
argus genome.
Estimation of C. argus genome size and sequencing
coverage
All the cleaned reads were subjected to 17-mer frequency dis-
tribution analysis [2]. As the total number of k-mers was about
5.90 × 10
10
and the peak of k-mers at a depth of 88, the genome
size of C. argus was calculated to be 670.4 Mb using the following
formula: genome size = k-mer
number / peak depth. Therefore,
the sequencing coverage was found to be ∼124.5 × based on the
estimated genome size.
De novo genome assembly and quality assessment
For whole genome assembly, SOAPdenovo2 [3] was used with
optimized parameters (-K 75) to construct contigs and original
scaffolds by using the reads from short-insert libraries. All reads
were then mapped onto contigs for scaffold construction by uti-
lizing the paired-end information of long-insert libraries. Some
intra-scaffold gaps were lled by local software using read-pairs
in which one end uniquely mapped to a contig and the other
end was located within a gap. Finally, a draft C. argus genome of
615.3 Mb was assembled, with a contig N50 size of 81.4 kb and a
scaffold N50 size of 4.5 Mb (Table 1).
Subsequently, the Core Eukaryotic Genes Mapping Approach
software [4] (version 2.3) with 248 conserved Core Eukaryotic
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Draft genome of the Northern snakehead 3
Table 1: summary of the Channa argus genome assembly and
annotation
Genome assembly
Contig N50 size (kb) 81
Contig number (>100 bp) 29 146
Scaffold N50 size (Mb) 4.5
Scaffold number (>100 bp) 5297
Total length (Mb) 615.3
Genome coverage (X) 224.6
The longest scaffold (bp) 18 736 006
Genome annotation
Protein-coding gene number 19 877
Mean transcript length (kb) 16.5
Mean exons per gene 10.5
Mean exon length (bp) 175.0
Mean intron length (bp) 1537.3
Genes was utilized to evaluate completeness of genes. Our
results demonstrated that the generated genome assembly
covered 242 of the 248 Core Eukaryotic Gene sequences, sug-
gesting a high level of completeness within the genome assem-
bly. Alongside this, we also used BUSCO (version 1.22) [5](the
representative vertebrate gene set containing 3023 single-copy
genes that are highly conserved in vertebrates) software to as-
sess the quality of the generated genome assembly. The assess-
ment demonstrated that the BUSCO value is 82.9%, containing
C: 66% [D: 1.4%], F: 16%, M: 17%, n: 3023 (C: complete [D: dupli-
cated], F: fragmented, M: missed, n: genes), suggesting a high
quality of the generated assembly.
Repeat sequence within the C. argus genome assembly
To analyze the C. argus genome, we employed Tandem Repeats
Finder [6] (version 4.04) with core parameters set as “Match = 2,
Mismatch = 7, Delta = 7, PM = 80, PI = 10, Minscore = 50, and
MaxPerid = 2000” to identify tandem repeats. Simultaneously,
RepeatModeler (version 1.04) and LTR
FINDER [7] were utilized
to construct a de novo repeat library with default parameters.
Subsequently, we used RepeatMasker [8] (version 3.2.9) to map
our assembled sequences on the Repbase TE (version 14.04) [9]
and the de novo repeat libraries to search for known and novel
transposable elements (TEs). In addition, the TE-related proteins
were annotated by using RepeatProteinMask software [8](ver-
sion 3.2.2). In summary, the total identied repeat sequences
accounted for 18.94% of the C. argus genome (Table 2). Among
them, long interspersed nuclear elements were the most abun-
dant type of repeat sequences and occupy 8.92% of the whole
genome.
Gene annotation
Gene annotation of the C. argus genome was conducted using
several approaches, including transcriptome-based prediction,
de novo prediction, and homology-based prediction. RNA-seq
datasets of pooled 13 tissues were obtained from our previous
work [10]. We mapped these RNA reads onto our genome assem-
bly using TopHat1.2 software [11], and then we employed Cuf-
inks (version 2.2.1) [12] to predict the gene structures. Further-
more, we performed Augustus (version 2.5.5) [13], GlimmerHMM
(version 3.0.1) [14], and GenScan (version 1.0) [15] softwares for
de novo prediction on the repeat-masked C. argus genome assem-
bly. The protein sequences of zebrash (Danio rerio)[16], Japanese
puffer (Fugu rubripes)[17], medaka (Oryzias latipes)[18], spotted
green puffersh (Tetraodon nigroviridis)[19] (the above 5 species
were downloaded from Ensembl release 75), blue spotted mud-
skipper (Boleophthalmus boddarti)[20], and golden arowana (Scle-
ropages formosus)[21] were mapped on the C. argus genome us-
ing TblastN with e-value ≤ 1e-5. Subsequently, Genewise2.2.0
software [22] was employed to predict the potential gene struc-
tures on all alignments. Finally, the above three datasets were
integrated to yield a comprehensive and nonredundant gene set
using GLEAN (https://sourceforge.net/projects/glean-gene/)[23]
with several lter steps (removing partial sequences or genes
shorter than 150 bp or prematurely terminated/frame-shifted
genes). The nal total gene set was composed of 19 877 genes,
with an average of 10.5 exons per gene (Table 1).
Construction of gene families and phylogenetic tree
We downloaded the protein sequences of zebrash [17],
Japanese puffer [18], stickleback (Gasterosteus aculeatus)[24],
spotted green puffersh [20], and medaka [19] from the Ensembl
Core database (release 75), and we also obtained the protein se-
quences of Asian seabass (Lates calcarifer)[25], blue spotted mud-
skipper [21], and golden arowana [22] from their correspond-
ing ftp websites, respectively. The consensus proteome set of
the above eight species and snakehead sh was ltered to re-
move those protein sequences <50 amino acids and resulted
in a dataset of 190 566 protein sequences, which was used as
the input le for OrthoMCL [26] to construct gene families. A to-
tal of 17 954 OrthoMCL families were built utilizing an effective
database size of 190 566 sequences for all-to-all BLASTP strategy
with an E-value of 1e-5 and a Markov Chain Clustering default
ination parameter. We further identied 24 gene families that
were specic in the snakehead sh (Fig. 2a).
Subsequently, we selected 1918 single-copy (only one gene
from each species) orthogroups from the above-mentioned 9
teleost species. We used MUSCLE (version 3.8.31) [27] to align the
Table 2: the detailed classication of repeat sequences of Channa argus
Repbase TEs TE protiens De novo Combined TEs
Type Length (bp) % in genome Length (bp) % in genome Length (bp) % in genome Length (bp) % in genome
DNA 17 984 515 2.92 6 784 728 1.10 25 663 752 4.17 35 435 946 5.76
LINE 16 799 343 2.73 17 563 763 2.85 54 890 557 8.92 60 651 866 9.86
SINE 4 512 385 0.73 0 0 6 672 552 1.08 9 026 285 1.47
LTR 4 421 728 0.72 3 031 607 0.49 24 144 657 3.92 26 983 318 4.39
Other 8125 0.001 0 0 0 0 8125 0.001
Unknown 0 0 0 0 9 413 375 1.53 9 413 375 1.53
Total 41 585 442 6.76 27 363 267 4.45 103 162 115 16.77 116 545 270 18.94
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4 Xu et al.
Figure: 2: genome evolution. (a) Orthologous gene families across ve sh genomes (Snakehead sh, Zebrash, Asian seabass, Mudskipper, and Arowana). (b) Phylogeny
of ray-nned shes (the arowana as the outgroup species).
protein sequences from the 1918 orthogroups, respectively. We
also converted protein alignments to their corresponding cod-
ing DNA sequence alignments using an in-house perl script. All
the translated coding DNA sequence sequences were then com-
bined into one “supergene” for each species. Nondegenerated
sites (4D) extracted from the supergenes were then joined into
new sequence of each species to construct a phylogenetic tree
(Fig. 2b) using MrBayes [28] (Version 3.2, with the GTR+gamma
model).
Conclusion
We report the rst whole genome sequencing, assembly, and
annotation of the Northern snakehead (Channa argus). The -
nal draft genome assembly is approximately 615.3 Mb, account-
ing for 91.8% of the estimated genome size (670.4 Mb). We
also predicted 19 877 protein-coding genes from the generated
assembly.
The draft genome assembly will be valuable resource for
genetic breeding, environmental DNA detection of invasive
species, and biological studies on this economically important
teleost sh. Based on these genomic data, researchers will be
able to develop genetic markers for further quantitative trait
locus and genome-wide association studies on growth traits.
These markers will also be very useful for DNA barcoding in
screening invasive C. argus for ecological protection.
Availability of supporting data
The raw sequencing reads of all libraries have been deposited at
NCBI (SRP078899). Further supporting data are available in the
GigaScience database, GigaDB [29].
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Draft genome of the Northern snakehead 5
Abbreviation
TE: transposable element.
Author contributions
PX designed the study. JX, CB, GL, JL, HD, YH, YX, and QS as-
sembled and annotated the genome. CB and YY performed the
evolution analysis. JX, YJ, XY, QL, and HZ analyzed the data. WP,
CD, SZ, and KC collected the sample and prepared the quality
control. JX, CB, QS, and PX wrote the manuscript. QS and PX par-
ticipated in discussions and provided advice. All authors read
and approved the nal manuscript.
Acknowledgements
This work was supported by Central Public-interest Scientic In-
stitution Basal Research Fund, CAFS (No. 2015C005, No. 2016HY-
JC0301), the National Natural Science Foundation of China (No.
31422057, No.31402291), the National Infrastructure of Fishery
Germplasm Resources of China (No. 2017DKA30470), Special
Project on the Integration of Industry, Education and Research
of Guangdong Province (No. 2013B090800017), Quality Inspec-
tion Programs of Scientic Research Project (No. 2015IK246), and
Shenzhen Special Program for Future Industrial Development
(No. JSGG20141020113728803).
Competing interests
The authors declare that they have no competing interests.
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