Skip to main content
Log in

RNA Sequencing to Study Gene Expression and SNP Variations Associated with Growth in Zebrafish Fed a Plant Protein-Based Diet

  • Original Article
  • Published:
Marine Biotechnology Aims and scope Submit manuscript

Abstract

The objectives of this study were to measure gene expression in zebrafish and then identify SNP to be used as potential markers in a growth association study. We developed an approach where muscle samples collected from low- and high-growth fish were analyzed using RNA-Sequencing (RNA-seq), and SNP were chosen from the genes that were differentially expressed between the low and high groups. A population of 24 families was fed a plant protein-based diet from the larval to adult stages. From a total of 440 males, 5 % of the fish from both tails of the weight gain distribution were selected. Total RNA was extracted from individual muscle of 8 low-growth and 8 high-growth fish. Two pooled RNA-Seq libraries were prepared for each phenotype using 4 fish per library. Libraries were sequenced using the Illumina GAII Sequencer and analyzed using the CLCBio genomic workbench software. One hundred and twenty-four genes were differentially expressed between phenotypes (p value < 0.05 and FDR < 0.2). From these genes, 164 SNP were selected and genotyped in 240 fish samples. Marker-trait analysis revealed 5 SNP associated with growth in key genes (Nars, Lmod2b, Cuzd1, Acta1b, and Plac8l1). These genes are good candidates for further growth studies in fish and to consider for identification of potential SNPs associated with different growth rates in response to a plant protein-based diet.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Subscribe and save

Springer+ Basic
EUR 32.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or Ebook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3

Similar content being viewed by others

References

  • Abdul Ajees A, Gunasekaran K, Volanakis JE, Narayana SVL, Kotwal GJ, Murthy HM (2006) The structure of complement C3b provides insights into complement activation and regulation. Nature 444:221–225

    Article  CAS  PubMed  Google Scholar 

  • Alami-Durante H, Médale F, Cluzeaud M, Kaushik SJ (2010) Skeletal muscle growth dynamics and expression of related genes in white and red muscles of rainbow trout fed diets with graded levels of a mixture of plant protein sources as substitutes for fishmeal. Aquaculture 303:50–58

    Article  CAS  Google Scholar 

  • Baranski M, Moen T, Vage D (2010) Mapping of quantitative trait loci for flesh colour and growth traits in Atlantic salmon (Salmo salar). Genet Sel Evol 42:17

    Article  PubMed Central  PubMed  Google Scholar 

  • Bostock J, McAndrew B, Richards R, Jauncey K, Telfer T, Lorenzen K, Little D, Ross L, Handisyde N, Gatward I, Corner R (2010) Aquaculture: global status and trends. Philos Trans R Soc Lond B Biol Sci 365:2897–2912

    Article  PubMed Central  PubMed  Google Scholar 

  • Bouza C, Hermida M, Pardo B, Vera M, Fernandez C, De La Herran R, Navajas-Perez R, Alvarez-Dios JA, Gomez-Tato A, Martinez P (2012) An Expressed Sequence Tag (EST)-enriched genetic map of turbot (Scophthalmus maximus): a useful framework for comparative genomics across model and farmed teleosts. BMC Genet 13:54

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Brand M, Granato M, Nüsslein-Volhard C (2002) Keeping and raising zebrafish. In: Nüsslein-Volhard C, Dahm R (eds) Zebrafish. Oxford University Press, Oxford

    Google Scholar 

  • Cánovas A, Rincon G, Islas-Trejo A, Wickramasinghe S, Medrano JF (2010) SNP discovery in the bovine milk transcriptome using RNA-Seq technology. Mamm Genome 21:592–598

    Article  PubMed Central  PubMed  Google Scholar 

  • Cánovas A, Rincon G, Islas-Trejo A, Jimenez-Flores R, Laubscher A, Medrano JF (2013) RNA sequencing to study gene expression and single nucleotide polymorphism variation associated with citrate content in cow milk. J Dairy Sci 96:2637–2648

    Article  PubMed  Google Scholar 

  • Castano-Sánchez C, Fuji K, Ozaki A, Hasegawa O, Sakamoto T, Morishima K, Nakayama I, Fujiwara A, Masaoka T, Okamoto H, Hayashida K, Tagami M, Kawai J, Hayashizaki Y, Okamoto N (2010) A second generation genetic linkage map of Japanese flounder (Paralichthys olivaceus). BMC Genomics 11:554

    Article  PubMed Central  PubMed  Google Scholar 

  • Conesa A, Götz S (2008) Blast2GO: a comprehensive suite for functional analysis in plant genomics. Int J Plant Genomics. doi: 10.1155/2008/619832

  • Conesa A, Götz S, García-Gómez JM, Terol J, Talón M, Robles M (2005) Blast2GO: a universal tool for annotation, visualization and analysis in functional genomics research. Bioinformatics 21:3674–3676

    Article  CAS  PubMed  Google Scholar 

  • Cui J, Liu S, Zahng B, Wang H, Sun H, Song S, Qui X, Liu Y, Wang X, Jiang Z, Liu Z (2014a) Transcriptome analysis of the gill and swimbladder of Takifugu rubripens by RNA-Seq. PLoS ONE 9:e85505

    Article  PubMed Central  PubMed  Google Scholar 

  • Cui J, Wang H, Liu S, Zhu L, Qiu X, Jiang Z, Wang X, Liu Z (2014b) SNP discovery from transcriptome of the swimbladder of Takifugu rubripes. PLoS ONE 9:e92502

    Article  PubMed Central  PubMed  Google Scholar 

  • FAO (2012) El estado mundial de la pesca y la acuicultura. Organización de las Naciones Unidas para la Alimentación y la Agricultura, Italy

    Google Scholar 

  • Fontaínhas-Fernandes A, Gomes E, Reis-Henriques MA, Coimbra J (1999) Replacement of fish meal by plant proteins in the diet of Nile tilapia: digestibility and growth performance. Aquac Int 7:57–67

    Article  Google Scholar 

  • Gómez-Requeni P, Mingarroa M, Calduch-Ginera J, Médale F, Martinc SAM, Houlihanc DF, Kaushik S, Pérez-Sánchez J (2004) Protein growth performance, amino acid utilisation and somatotropic axis responsiveness to fish meal replacement by plant protein sources in gilthead sea bream (Sparus aurata). Aquaculture 232:493–510

    Article  Google Scholar 

  • Guyon R, Senger F, Rakotomanga M, Sadequi N, Volckaert FAM, Hitte C, Galibert F (2010) A radiation hybrid map of the European sea bass (Dicentrarchus labrax) based on 1581 markers: synteny analysis with model fish genomes. Genomics 96:228–238

    Article  CAS  PubMed  Google Scholar 

  • Hardy RW (2010) Utilization of plant proteins in fish diets: effects of global demand and supplies of fishmeal. Aquac Res 41:770–776

    Article  CAS  Google Scholar 

  • Hedrera M, Galdames J, Gimenez-Reyes M, Reyes A, Avandaño-Herrera R, Romero J, Feijoo CG (2013) Soybean meal induces intestinal inflammation in zebrafish larvae. PLoS ONE 8:e69983

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Houston RD, Taggart JB, Cezard T, Bekaert M, Lowe NR, Downing A, Talbot R, Bishop SC, Archibald AL, Bron JE, Penman DJ, Davassi A, Brew F, Tinch A, Gharbi K, Hamilton A (2014) Development and validation of a high density SNP genotyping array for Atlantic salmon (Salmo salar). BMC Genomics 15:90

    Article  PubMed Central  PubMed  Google Scholar 

  • Hutson AM, Liu Z, Kucuktas H, Umali-Maceina G, Su B, Dunham RA (2014) Quantitative trait loci map for growth and morphometric traits using a channel catfish x blue catfish interspecific hybrid system. J Anim Sci 92:1850–1865

    Article  CAS  PubMed  Google Scholar 

  • Ibba M, Söll D (2000) Aminoacyl-tRNA synthesis. Annu Rev Biochem 69:617–650

    Article  CAS  PubMed  Google Scholar 

  • Jiang Y, Gao X, Liu S, Zhang Y, Liu H, Sun F, Bao L, Waldbieser G, Liu Z (2013) Whole genome comparative analysis of channel catfish (Ictalurus punctatus) with four model fish species. BMC Genomics 14:780

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Jimenez-Preitner M, Berney X, Uldry M, Vitali A, Cinti S, Ledford JG, Thorens B (2011) Plac8 is an inducer of C/EBPβ required for brown fat differentiation, thermoregulation, and control of body weight. Cell Metab 14:658–670

    Article  CAS  PubMed  Google Scholar 

  • Küttner E, Moghadam H, Skúlason S, Danzmann R, Ferguson M (2011) Genetic architecture of body weight, condition factor and age of sexual maturation in Icelandic Arctic charr (Salvelinus alpinus). Mol Genet Genomics 286:67–79

    Article  PubMed  Google Scholar 

  • Laing NG, Dye DE, Wallgren-Pettersson C, Richard G, Monnier N, Lillis S, Winder TL, Lochmüller H, Graziano C, Mitrani-Rosenbaum S, Twomey D, Sparrow JC, Beggs AH, Nowak KJ (2009) Mutations and polymorphisms of the skeletal muscle α-actin gene (ACTA1). Hum Mutat 30:1267–1277

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Ledford JG, Kovarova M, Koller BH (2007) Impaired host defense in mice lacking ONZIN. J Immunol 178:5132–5143

    Article  CAS  PubMed  Google Scholar 

  • Liu S, Gao G, Palti Y, Cleveland BM, Weber GM, Rexroad CE III (2014a) RNA-Seq analysis of early hepatic response to handling and confinement stress in rainbow trout. PLoS ONE 2:e88492

    Article  Google Scholar 

  • Liu S, Sun L, Li Y, Sun F, Jiang Y, Zhang Y, Zhang J, Feng J, Kaltenboeck L, Kucuktas H, Liu Z (2014b) Development of the catfish 250K SNP array for genome-wide association studies. BMC Res Notes 7:135

    Article  PubMed Central  PubMed  Google Scholar 

  • Long Y, Song G, Yan J, He X, Li Q, Cui Z (2013) Transcriptomic characterization of cold acclimation in larval zebrafish. BMC Genomics 14:612

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Médale F, Boujard T, Vallee F, Blanc D, Mambrini M, Roem R, Kaushik S (1998) Voluntary feed intake, nitrogen and phosphorus losses in rainbow trout (Oncorhynchus mykiss) fed increasing dietary levels of soy protein concentrate. Aquat Living Resour 11(4):239–246

    Article  Google Scholar 

  • Mignone F, Gissi C, Liuni S, Pesole G (2002) Untranslated regions of mRNAs. Genome Biol 3(3): reviews0004.1–0004.10

  • Morais S, Pratoomyot J, Taggart J, Bron J, Guy D, Bell J, Tocher D (2011) Genotype-specific responses in Atlantic salmon (Salmo salar) subject to dietary. BMC Genomics 12:255

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Mortazavi A, Williams BA, Mccue K, Schaeffer L, Wold B (2008) Mapping and quantifying mammalian transcriptomes by RNA-Seq. Nat Methods 5:621–628

    Article  CAS  PubMed  Google Scholar 

  • Mundheim H, Aksnes A, Hope B (2004) Growth, feed efficiency and digestibility in salmon (Salmo salar L.) fed different dietary proportions of vegetable protein sources in combination with two fish meal qualities. Aquaculture 237:315–331

    Article  Google Scholar 

  • Mutch DM, Wahli W, Williamson G (2005) Nutrigenomics and nutrigenetics: the emerging faces of nutrition. FASEB J 19:1602–1616

    Article  CAS  PubMed  Google Scholar 

  • Naylor RL, Hardy RW, Bureau DP, Chiu A, Elliott M, Farrell AP, Forster I, Gatlin DM, Goldburg RJ, Hua K, Nichols PD (2009) Feeding aquaculture in an era of finite resources. Proc Natl Acad Sci 106:15103–15110

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • NRC (1993) In: Press NA (Ed) Nutrient requirements of fish. Washington DC, USA

  • Perry SE, Robinson P, Melcher A, Quirke P, Bühring HJ, Cook GP, Blair GE (2007) Expression of the CUB domain containing protein 1 (CDCP1) gene in colorectal tumour cells. FEBS Lett 581:1137–1142

    Article  CAS  PubMed  Google Scholar 

  • Pongmaneerat J, Watanabe T, Takeuchi T, Satoh T (1993) Use of different protein meals as partial or total substitution for fish meal in carp diets. Nippon Suisan Gakkaishi 59:1249–1257

    Article  CAS  Google Scholar 

  • Price AL, Patterson NJ, Plenge RM, Weinblatt ME, Shadick NA, Reich D (2006) Principal components analysis corrects for stratification in genome-wide association studies. Nat Genet 38:904–909

    Article  CAS  PubMed  Google Scholar 

  • Qian X, Ba Y, Zhuang Q, Zhong G (2014) RNA-seq technology and its application in fish transcriptomics. OMICS J Integr Biol 18(2):98–110

    Article  CAS  Google Scholar 

  • Rincon G, Farber EA, Farber CR, Nkrumah JD, Medrano JF (2009a) Polymorphisms in the STAT6 gene and their association with carcass traits in feedlot cattle. Anim Genet 40:878–882

    Article  CAS  PubMed  Google Scholar 

  • Rincon G, Islas-Trejo A, Casellas J, Ronin Y, Soller M, Lipkin E, Medrano JF (2009b) Fine mapping and association analysis of a quantitative trait locus for milk production traits on Bos taurus autosome 4. J Dairy Sci 92:758–764

    Article  CAS  PubMed  Google Scholar 

  • Rincon G, Islas-Trejo A, Castillo AR, Bauman DE, German BJ, Medrano JF (2012) Polymorphisms in genes in the SREBP1 signalling pathway and SCD are associated with milk fatty acid composition in Holstein cattle. J Dairy Res 79:66–75

    Article  CAS  PubMed  Google Scholar 

  • Salem M, Vallejo RL, Leeds TD, Palti Y, Liu S, Sabbagh A, Rexroad CE, Yao J (2012) RNA-seq identifies SNP markers for growth traits in rainbow trout. PLoS One 7(5):e36264

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Skwarek-Maruszewska A, Boczkowska M, Zajac AL, Kremneva E, Svitkina T, Dominguez R, Lappalainen P (2010) Different localizations and cellular behaviors of leiomodin and tropomodulin in mature cardiomyocyte sarcomeres. Mol Biol Cell 21:3352–3361

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Storey J (2002) A direct approach to false discovery rates. J R Stat Soc Se B 64:479–498

    Article  Google Scholar 

  • Takebayashi H, Yamamoto N, Umino A, Nishikawa T (2009) Developmentally regulated and thalamus-selective induction of leiomodin2 gene by a schizophrenomimetic, phencyclidine, in the rat. Int J Neuropsychopharmacol 12(8):1111–1126

    Article  CAS  PubMed  Google Scholar 

  • Tave D (1993) Genetics for fish hatchery managers. Van Nostrand Reinhold, New York

    Google Scholar 

  • Ulloa PE, Iturra P, Neira R, Araneda C (2011) Zebrafish as a model organism for nutrition and growth: towards comparative studies of nutritional genomics applied to aquacultured fishes. Rev Fish Biol Fish 21:649–666

  • Ulloa PE, Peña A, Lizama CD, Araneda C, Iturra P, Neira R, Medrano JF (2013) Growth response and expression of muscle growth-related candidate genes in adult zebrafish fed plant and fishmeal protein-based diets. Zebrafish 10:1

    Article  Google Scholar 

  • Ulloa PE, Medrano JF, Feijoo CG (2014) Zebrafish as animal model for aquaculture nutrition research. Front Genet 5:313

    Article  PubMed Central  PubMed  Google Scholar 

  • Vilhelmsson OT, Martin SAM, Médale F, Kaushik SJ, Houlihan DF (2004) Dietary plant-protein substitution affects hepatic metabolism in rainbow trout (Oncorhynchus mykiss). Brit J Nutr 92:71–80

    Article  CAS  PubMed  Google Scholar 

  • Von Hertell U, Hörstgen-Schwark G, Langholz HJ, Jung B (1990) Family studies on genetic variability in growth and reproductive performance between and within test fish populations of the zebrafish, Brachydanio rerio. Aquaculture 85:307–315

    Article  Google Scholar 

  • Weatherley A, Gill H (1987) The biology of fish growth. Academic press, London, p 443

  • Wickramasinghe S, Cánovas A, Rincón G, Medrano JF (2014) RNA-sequencing: a tool to explore new frontiers in animal genetics. Livest Prod Sci. doi: 10.1016/j.livsci.2014.06.015i

  • Wright D, Nakamichi R, Krause J, Butlin R (2006) QTL analysis of behavioral and morphological differentiation between wild and laboratory zebrafish (Danio rerio). Behav Genet 36:271–284

    Article  PubMed  Google Scholar 

  • Wringe B, Devlin R, Ferguson M, Moghadam H, Sakhrani D, Danzmann R (2010) Growth-related quantitative trait loci in domestic and wild rainbow trout (Oncorhynchus mykiss). BMC Genet 11:63

    Article  PubMed Central  PubMed  Google Scholar 

  • Xia J, Liu F, Zhu Z, Fu J, Feng J, Li J, Yue G (2010) A consensus linkage map of the grass carp (Ctenopharyngodon idella) based on microsatellites and SNPs. BMC Genomics 11:135

    Article  PubMed Central  PubMed  Google Scholar 

Download references

Acknowledgments

PEU wishes to thank the Animal Genomic Laboratory research group at the University of California-Davis where the RNA-Seq experiments were performed and the UC-Davis Genome Center for excellent technical expertise in performing the Illumina GAII sequencing. PEU received a doctoral fellowship from the Consorcio Empresarial de Genética y Desarrollo Biotecnológico para la Industria Salmonera (Aquainnovo S.A.), and Apoyo a la Realización de Tesis Doctoral Fellowship from Conicyt AT-24091052, and Becas Chile Scholarship 75110126. PEU wishes to thank the Programa Convenio Desempeño EVOLUCIONA de la Universidad de la Frontera and its Programa de Doctorado en Ciencias de Recursos Naturales.

A special thanks goes to Kamila Solis for helping in the improve the quality of the Fig. 3.

Conflict of Interest

The authors declare that they have no competing interests.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Pilar E. Ulloa.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary file 1

(DOCX 22 kb)

Supplementary file 2

(DOCX 26 kb)

Supplementary file 3

(DOCX 22 kb)

Supplementary file 4

(DOCX 23 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Ulloa, P.E., Rincón, G., Islas-Trejo, A. et al. RNA Sequencing to Study Gene Expression and SNP Variations Associated with Growth in Zebrafish Fed a Plant Protein-Based Diet. Mar Biotechnol 17, 353–363 (2015). https://doi.org/10.1007/s10126-015-9624-1

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10126-015-9624-1

Keywords

Navigation