Ciona: A Model for Developmental Genomics


The ascidian Ciona intestinalis is an excellent model system for studying developmental biology due to the simplicity of its embryogenesis and genomics. The Ciona tadpole larva is composed of only 2600 cells and possesses the basic body plan of chordates. The 117‐Mbp‐long euchromatic genome encodes approximately 320 core transcription factors and 125 major signal transduction molecules, the expression profiles of which have been described at the single‐cell level in Ciona embryos. Not only the function of developmentally relevant genes can be explored by means of suppressed and/or ectopic expression, but also a convenient electroporation method of reporter gene constructs has been developed to identify cis‐regulatory modules. High throughput and genome‐wide approaches can be used to uncover gene regulatory networks involved in cell specification and differentiation. Transposon‐mediated transgenesis also provides a new strategy for elucidating the mechanisms of gene expression and function. These and other advantages of the Ciona system recommend future developmental biology studies with this model.

Key Concepts:

  • The Ciona tadpole larva represents the basic and most simplified chordate body plan, and its embryogenesis is well characterised.

  • The Ciona intestinalis genome has been decoded. The 117‐Mbp euchromatic genome, which is packaged into 14 pairs of chromosomes, contains 15 254 genes including ∼320 core TF genes and 125 major signalling molecule genes.

  • Most of these developmentally relevant genes have been chromosomally mapped.

  • The spatial and temporal expression profiles of these developmentally relevant genes have been elucidated.

  • Gene function can be easily studied by suppressed and/or ectopic expression.

  • A convenient electroporation method allows quantitative and qualitative examination of cis‐regulatory modules responsible for gene expression.

  • The regulatory networks of TF and signalling molecule genes that establish the blueprint of the chordate body plan have been uncovered.

  • The short generation time allows forward genetics of Ciona, a first for a marine invertebrate.

  • Transgenic lines have been established using Minos transposons to reveal gene function and expression.

Keywords: ascidian; Ciona intestinalis; tadpole larvae; basic chordate body plan; decoded genome; transcriptomics; forward genetics; transcription factors; cell–cell signalling molecules; cis‐regulatory modules; gene regulatory network

Figure 1.

The ascidian urochordate C. intestinalis. (a) Adults with incurrent and outcurrent syphons. The white duct is the sperm duct and the parallel orange duct is the egg duct. (b–g) Embryogenesis. Embryos were dechorionated to show their outer morphology clearly: (b) Fertilised egg, (c) 16‐cell embryo, (d) gastrula (approximately 150 cells), (e, f) tailbud embryos and (g) tadpole larva. (h) A juvenile, a few days after metamorphosis, showing the internal structures: ds, digestive system; en, endostyle: ht, heart; os, neuronal complex; and pg, pharyngeal gill. (i) Fate map of the 110‐cell embryo, animal hemisphere (left) and vegetal hemisphere (right). Blastomeres are named according to Conklin's nomenclature and coloured according to developmental fate restriction. Green, epidermis; dark blue, brain; light blue, nerve cord; magenta, notochord; light blue, trunk lateral cells; dark green, mesenchyme; yellow, endoderm; and red, muscle. (j) Oblique lateral confocal section through a mid‐tailbud stage C. savignyi embryo. The 40 notochord cells (green) were marked with a stable Brachyury:GFP transgene. Cell peripheries were labelled with phalloidin and manually pseudocoloured to show the endoderm (blue), muscle (red), neural tube (yellow) and epidermis (magenta). CNS, central nervous system; End, endoderm; Epi, epidermis; NC, nerve cord; Mch, mesenchyme; Mu, muscle; Not, notochord. Reproduced with permission from ‘Development’ 1 January 2008, 135 (1) (cover image) Image: William Smith.

Figure 2.

Chromosomal map of 373 core TF genes and 111 major cell signalling molecule genes in C. intestinalis. Families of TF are shown by different coloured discs, whereas families of cell signalling molecules are shown by different coloured arrowheads (bottom right). Centromeric regions are shown by dark blue dashed lines. Red dashed lines indicate three rDNA cluster regions and green dashed lines indicate a histone cluster region. Blue dashed lines indicate unmapped regions. The left and right vertical lines of each chromosome indicate the 5′–3′ and 3′–5′ alignment, respectively. The telomeric regions on the short arms of chromosomes 12, 13 and 14 are ordered arbitrarily.

Figure 3.

A method to examine regulatory modules responsible for specific gene expression by electroporation of reporter gene constructs. (a) Fertilised or unfertilised eggs (brown circles) within an electroporation chamber filled with seawater containing reporter gene constructs (black bars). (b) Culture of manipulated embryos in agar‐coated plastic dishes. (c) Detection of reporter (lacZ or GFP) expression in larvae. In this case, GFP expression driven by Brachyury promoter is evident. This process (from a to c) only takes ∼20 h. Reproduced with permission from: Cover image from Science vol. 298, no. 5601, 13 December 2002. Image: Mei Wang. © American Association for the Advancement of Science.

Figure 4.

Regulatory network of zygotically expressed TF and STM genes in the early Ciona embryo. (a) A 64‐cell stage embryo viewed from the animal (upper right) or vegetal (lower right) pole, with nomenclature of blastomeres, and the line of embryonic cells (left) indicated. (b) Expression patterns of 36 regulatory genes zygotically expressed at the 64‐cell stage. (c) The regulatory network of 76 genes expressed in early embryos, up to the early gastrula stage. The genes are ordered alphabetically from the top‐left corner to the bottom‐right corner. The chromosomal localisation of the genes is also shown in light boxes, with short (left) and long (right) arms. Lines show the gene regulatory networks, dark blue lines and arrows indicate the same chromosomal interactions, orange lines show interchromosomal interactions and green arrows indicate autoregulatory interactions.

Figure 5.

Transgenic Ciona lines. (a) Production of transgenic lines with specific markers. Unfertilised eggs are first microinjected with transposon vector including GFP and transposase mRNA. They are then inseminated and allowed to develop into mature adults. Founder sperm is used to fertilise eggs of wild‐type animals. These eggs are then cultured to larval, juvenile and adult stages, from which specimens with GFP expression are selected. (b–f) Examples of transgenic lines with specific markers: (b) A transgenic line in which GFP is specifically expressed in the epidermis, (c) notochord, (d) muscle or (e) central nervous system of larva. (f) A line in which juvenile expresses GFP in the nervous system and red fluorescent protein in the endostyle. Courtesy of Dr. Sasakura.



Azumi K, Sabau SV, Fujie M et al. (2007) Gene expression profile during the life cycle of the urochordate Ciona intestinalis. Developmental Biology 308: 572–582.

Christiaen L, Davidson B, Kawashima T et al. (2008) The transcription/migration interface in heart precursors of Ciona intestinalis. Science 320: 1349–1352.

Cloney L (1978) Ascidian metamophosis; review and analysis. In: Chia F‐S and Rice ME (eds) Settlement and Metamorphosis of Marine Invertebrate Larvae, pp. 255–282. Amsterdam: Elsevier.

Conklin EG (1905) The organization and cell lineage of the ascidian egg. Journal of the Academy of Natural Sciences of Philadelphia 13: 1–119.

Corbo JC, Levine M and Zeller RW (1997) Characterization of a notochord‐specific enhancer from the Brachyury promoter region of the ascidian, Ciona intestinalis. Development 124: 589–602.

Davidson B (2007) Ciona intestinalis as a model for cardiac development. Seminars in Cell and Developmental Biology 18: 16–26.

Davidson B, Shi W, Beb J, Christiaen L and Levine M (2006) FGF signaling delineates the cardiac progenitor field in the simple chordate, Ciona intestinalis. Genes and Development 20: 2728–2738.

Davidson EH (2006) The Regulatory Genome. Gene Regulatory Networks in Development and Evolution. San Diego, CA: Academic Press.

Dehal P, Satou Y, Campbell RK et al. (2002) The draft genome of Ciona intestinalis: insights into chordate and vertebrate origins. Science 298: 2157–2167.

Di Gregorio A and Levine M (2002) Analyzing gene regulation in ascidian embryos: new tools for new perspectives. Differentiation 70: 132–139.

Gilbert S (2010) Developmental Biology, 9th edn. Sunderland, MA: Sinauer Associates, Inc.

Hamada M, Shimozono N, Ohta N et al. (2011) Expression of neuropeptide‐ and hormone‐encoding genes in the Ciona intestinalis larval brain. Developmental Biology 352: 202–214.

Hirano T and Nishida H (1997) Developmental fates of larval tissues after metamorphosis in ascidian Halocynthia roretzi. I. Origin of mesodermal tissues of the juvenile. Developmental Biology 192: 199–210.

Horie T, Shinki R, Ogura Y et al. (2011) Ependymal cells of chordate larvae are stem‐like cells that form the adult nervous system. Nature 469: 525–528.

Hotta K, Takahashi H, Satoh N and Gojobori T (2008) Brachyury‐downstream gene sets in a chordate, Ciona intestinalis: integrating notochord specification, morphogenesis and chordate evolution. Evolution and Development 10: 37–51.

Ikuta T, Satoh N and Saiga H (2010) Limited functions of Hox genes in the larval development of the ascidian Ciona intestinalis. Development 137: 1505–1513.

Ikuta T, Yoshida N, Satoh N and Saiga H (2004) Ciona intestinalis Hox gene cluster: its dispersed structure and residual colinear expression in development. Proceedings of the National Academy of Sciences of the USA 101: 15118–15123.

Imai KS, Hino K, Yagi K, Satoh N and Satou Y (2004) Gene expression profiles of transcription factors and signaling molecules in the ascidian embryo: towards a comprehensive understanding of gene networks. Development 131: 4047–4058.

Imai KS, Levine M, Satoh N and Satou Y (2006) Regulatory blueprint for a chordate embryo. Science 312: 1183–1187.

Imai KS, Stolfi A, Levine M and Satou Y (2009) Gene regulatory networks underlying the compartmentalization of the Ciona central nervous system. Development 136: 285–293.

Johnson DS, Davidson B, Brown CD et al. (2004) Noncoding regulatory sequences of Ciona exhibit strong correspondence between evolutionary constraint and functional importance. Genome Research 14: 2448–2456.

Joly JS, Kano S, Matsuoka T et al. (2007) Culture of Ciona intestinalis in closed systems. Developmental Dynamics 236: 1832–1840.

Keys DN, Lee BI, Di Gregorio A et al. (2005) A saturation screen for cis‐acting regulatory DNA in the Hox genes of Ciona intestinalis. Proceedings of the National Academy of Sciences of the USA 102: 679–683.

Khoueiry P, Rothbacher U, Ohtsuka Y et al. (2010) A cis‐regulatory signature in ascidians and flies, independent of transcription factor binding sites. Current Biology 20: 792–802.

Kumano G and Nishida H (2007) Ascidian embryonic development: an emerging model system for the study of cell fate specification in chordates. Developmental Dynamics 236: 1732–1746.

Kusakabe T, Yoshida R, Ikeda Y and Tsuda M (2004) Computational discovery of DNA motifs associated with cell if type‐specfic gene expression in Ciona. Developmental Biology 276: 563–580.

Lemaire P (2009) Unfolding a chordate developmental program, one cell at a time: invariant cell lineages, short‐range inductions and evolutionary plasticity in ascidians. Developmental Biology 332: 48–60.

Nakatani Y and Nishida H (1994) Induction of notochord during ascidian embryogenesis. Developmental Biology 166: 289–299.

Nakatani Y, Moody R and Smith WC (1999) Mutations affecting tail and notochord development in the ascidian Ciona savignyi. Development 126: 3293–3301.

Nishida H (1987) Cell lineage analysis in ascidian embryos by intracellular injection of a tracer enzyme III. Up to the tissue restricted stage. Developmental Biology 121: 526–541.

Nishida H (2005) Specification of embryonic axis and mosaic development in ascidians. Developmental Dynamics 233: 1177–1193.

Nishiyama A and Fujiwara S (2008) RNA interference by expressing short hairpin RNA in the Ciona intestinalis embryo. Development, Growth and Differentiation 50: 521–529.

Reverberi G (1971) Ascidians. In: Reverberi G (ed.) Experimental Embryology of Marine and Fresh‐water Invertebrates, pp. 507–550. Amsterdam: North Holland.

Rosner A, Rabinowitz C, Moiseeva E et al. (2007) BS‐Cadherin in the colonial urochordate Botryllus schlosseri: one protein, many functions. Developmental Biology 304: 687–700.

Sasakura Y (2007) Germline transgenesis and insertional mutagenesis in the ascidian Ciona intestinalis. Developmental Dynamics 236: 1758–1767.

Sasakura Y, Awazu S, Chiba S and Satoh N (2003) Germ‐line transgenesis of the Tc1/mariner superfamily transposon Minos in Ciona intestinalis. Proceedings of the National Academy of Sciences of the USA 100: 7726–7730.

Sasakura Y, Kanda M, Ikeda T et al. (2012) Retinoic acid‐driven Hox1 is required in the epidermis for forming the otic/atrial placodes during ascidian metamorphosis. Development 139: 2156–2160.

Sasakura Y, Nakashima K, Awazu S et al. (2005) Trnsposon‐mediated insertional mutagenesis revealed the functions of animal cellulose synthase in the ascidian Ciona intestinalis. Proceedings of the National Academy of Sciences of the USA 102: 15134–15139.

Satoh N (1994) Developmental Biology of Ascidians. Cambridge: Cambridge University Press.

Satoh N, Satou Y, Davidson B and Levine M (2003) Ciona intestinalis: an emerging model for whole‐genome analyses. Trends in Genetics 19: 376–381.

Satou Y and Satoh N (1996) Two cis‐regulatory elements are essential for the muscle‐specific expression of an actin gene in the ascidian embryo. Development Growth & Differentiation 38: 565–573.

Satou Y and Satoh N (2003) Genomewide surveys of developmentally relevant genes in Ciona intestinalis. Development Genes and Evolution 213: 211–212.

Satou Y, Mineta K, Ogasawara M et al. (2008) Improved genome assembly and evidence‐based global gene model set for the chordate Ciona intestinalis: new insight into intron and operon populations. Genome Biology 9: R152.

Shi W, Levine M and Davidson B (2005) Unraveling genomic regulatory networks in the simple chordate, Ciona intestinalis. Genome Research 15: 1668–1674.

Shoguchi E, Hamaguchi M and Satoh N (2008) Genome‐wide network of regulatory genes for construction of a chordate embryo. Developmental Biology 316: 498–509.

Shoguchi E, Kawashima T, Satou Y et al. (2006) Chromosomal mapping of 170 BAC clones in the ascidian Ciona intestinalis. Genome Research 16: 297–303.

Small KS, Brudno M, Hill MM and Sidow A (2007) A haplome alignment and reference sequence of the highly polymorphic Ciona savignyi genome. Genome Biology 8: R41.

Tassy O, Dauga D, Daian F et al. (2010) The ANISEED database: digital representation, formalization, and elucidation of a chordate developmental program. Genome Research 20: 1459–1468.

Tsagkogeorga G, Cahais V and Galtier N (2012) The population genomics of a faster evolver: high level of diversity, functional constrain and molecular adaptation in the tunic Ciona intestinalis. Genome Biology and Evolution 4: 852–861. doi:10.1093/gbe/evs054.

Yamada L, Kobayashi K, Satou Y and Satoh N (2005) Microarray analysis of localization of maternal transcripts in eggs and early embryos of the ascidian, Ciona intestinalis. Developmental Biology 284: 536–550.

Yamada L, Shoguchi E, Wada S et al. (2003) Morpholino‐based gene knockdown screen of novel genes with developmental function in Ciona intestinalis. Development 130: 6485–6495.

Yang J‐H, Shao P, Zhou H, Chen Y‐Q and Qu L‐H (2010) deepBase: a database for deeply annotating and mining deep sequencing data. Nucleic Acids Research 38: D123–D130.

Further Reading

Butts T, Holland PW and Ferrier DE (2008) The urbilaterian Super‐Hox cluster. Trends in Genetics 24: 259–262.

Jeffery WR and Swalla BJ (1997) Tunicates. In: Gilbert SF and Raunio AM (eds) Embryology, pp. 331–364. Sunderland, MA: Sinauer.

Meinertzhagen IA, Lemaire P and Okamura Y (2004) The neurobiology of the ascidian tadpole larva: recent development in an ancient chordate. Annual Review of Neurosciences 27: 453–458.

Satoh N (2001) Ascidian embryos as a model system to analyze expression and function of developmental genes. Differentiation 68: 1–12.

Satoh N (2008) An aboral‐dorsalization hypothesis for chordate origin. Genesis 46: 614–622.

Satou Y, Satoh N and Imai KS (2009) Gene regulatory networks in the early ascidian embryo. Biochimica et Biophysica Acta 1789: 268–273.

Contact Editor close
Submit a note to the editor about this article by filling in the form below.

* Required Field

How to Cite close
Satoh, Noriyuki(Mar 2013) Ciona: A Model for Developmental Genomics. In: eLS. John Wiley & Sons Ltd, Chichester. [doi: 10.1002/9780470015902.a0021411]