Research advances in the genomics and applications for molecular breeding of aquaculture animals

Effects of single or conjoint administration of lactic acid bacteria as potential probiotics on growth, immune response and disease resistance of snakehead fish (Channa argus).

// Mi Ou 1, 2 , Cheng Yang 1, 2 , Qing Luo 3 , Rong Huang 1 , Ai-Di Zhang 1 , Lan-Jie Liao 1 , Yong-Ming Li 1 , Li-Bo He 1 , Zuo-Yan Zhu 1 , Kun-Ci Chen 3 and Ya-Ping Wang 1 1 State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072, China 2 University of Chinese Academy of Sciences, Beijing 100049, China 3 Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou 510380, China Correspondence to: Ya-Ping Wang, email: wangyp@ihb.ac.cn Kun-Ci Chen, email: chenkunci@aliyun.com Keywords: next generation sequencing; sex-specific molecular markers; snakehead Received: April 19, 2017     Accepted: August 07, 2017     Published: October 19, 2017 ABSTRACT We described a next generation sequencing (NGS)-based approach to identify sex-specific markers and subsequently determine whether a species has male or female heterogamety. To test the accuracy of this technique, we examined the snakehead ( Channa argus ), which is economically important freshwater fish in China. Males grow faster than females, and there is significant interest in developing methods to skew breeding towards all-males to increase biomass yields. NGS was conducted on DNAs of individual female and male, the male reads were spitted into 60 bp K-mers and aligned to the female reference genome assembled by female reads, unaligned male K-mers-60 were kept in next filter process. Meanwhile, DNA sample of 48 females was pooled and sequenced, this data was further used to filter out the previous unaligned male K-mers-60. Hence, numbers of candidate Y chromosome-specific sequences were screened out, their sex-specificity were validated in wild snakeheads through PCR amplification. Finally, three Y chromosome-specific fragments (Contig-275834, Contig-359642, and Contig-418354) were identified, and specific primers were obtained to distinguish the sex of snakehead. Additionally, a pair of primers of Contig-275834 (275834X/Y-F and 275834X/Y-R) was exploited to distinguish XX females, XY males, and YY super-males, whose amplification products of different lengths were produced for different sexes. Therefore, our work demonstrated the ability of NGS data in identification of sex-specific markers, and the pipeline adopted in our study could be applied in any species of sex differentiation. Furthermore, the sex-specific markers have tremendous potential for improving the efficiency of all-male breeding practices in snakehead.

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An NGS-based approach for the identification of sex-specific markers in snakehead (Channa argus).

Transcriptome analysis of immune-related gene expression in hybrid snakehead (Channa maculata ♀ × Channa argus ♂) after challenge with Nocardia seriolae.

Detecting and documenting the occurrence of invasive species outside their native range requires tools to support their identification. This can be challenging for taxa with diverse life stages and/or problematic or unresolved morphological taxonomies. DNA barcoding provides a potent method for identifying invasive species, as it allows for species identification at all life stages, including fragmentary remains. It also provides an efficient interim taxonomic framework for quantifying cryptic genetic diversity by parsing barcode sequences into discontinuous haplogroup clusters (typical of reproductively isolated species) and labelling them with unique alphanumeric identifiers. Snakehead fishes are a diverse group of opportunistic predators endemic to Asia and Africa that may potentially pose significant threats as aquatic invasive species. At least three snakehead species (Channa argus, C. maculata, and C. marulius) are thought to have entered North America through the aquarium and live-food fish markets, and have established populations, yet their origins remain unclear. The objectives of this study were to assemble a library of DNA barcode sequences derived from expert identified reference specimens in order to determine the identity and aid invasion pathway analysis of the non-indigenous species found in North America using DNA barcodes. Sequences were obtained from 121 tissue samples representing 25 species and combined with public records from GenBank for a total of 36 putative species, which then partitioned into 49 discrete haplogroups. Multiple divergent clusters were observed within C. gachua, C. marulius, C. punctata and C. striata suggesting the potential presence of cryptic species diversity within these lineages. Our findings demonstrate that DNA barcoding is a valuable tool for species identification in challenging and under-studied taxonomic groups such as snakeheads, and provides a useful framework for inferring invasion pathway analysis.

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Calibrating snakehead diversity with DNA barcodes: expanding taxonomic coverage to enable identification of potential and established invasive species.

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.

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Draft genome of the Northern snakehead, Channa argus

This paper reports the complete mitochondrial genome (mitogenome) sequence of Channa maculate (Taiwan snakehead). The mitogenome of Taiwan snakehead was determined to be 16,558 bp in length. It contains 13 typical vertebrate protein-coding genes, 22 tRNA genes, 2 rRNA genes, 1 OL (putative light-strand origin of replication), and 1 putative control region (D-loop). This is the first report on the complete mitogenome sequence of Channa maculata.

The complete mitochondrial genome of Channa maculata (Perciformes: Channidae)

The invention discloses a primer pair and kit for distinguishing Channa argus and Channa maculata, and a PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) detection method. The primer pair comprises a forward primer and a reverse primer, wherein the polynucleotide sequence of the forward primer is disclosed as SEQ ID NO.1, and the polynucleotide sequence of the reverse primer is disclosed as SEQ ID NO.2. The kit containing the primer pair also comprises a PCR conventional reagent, a Taq DNA (deoxyribonucleic acid) polymerase, a 100*PCR buffer, dNTPs (deoxyribonucleotide triphosphates), a digestion reagent, a 10*buffer, a digestion buffer and an EcoRI restriction enzyme. The invention also discloses a PCR-RFLP detection method for distinguishing Channa argus and Channa maculata by using the primer pair and the kit. The method disclosed by the invention is simple to operate, only needs to cut a small amount of fin rays or muscles on the premise of ensuring the survival of the fish species, quickly and accurately identifies the fish species, enhances the accuracy and reliability of the identification result, and greatly enhances the work efficiency.

Primer pair and kit for distinguishing Channa argus and Channa maculata, and PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) detection method

Kunci Chen

Papers

Draft genome of the Northern snakehead, Channa argus

The complete mitochondrial genome of Channa maculata (Perciformes: Channidae)

Primer pair and kit for distinguishing Channa argus and Channa maculata, and PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) detection method