Oocytes of a single individual of Ciona intestinalis (Roscoff, France) were fertilized by a mixture of sperm of two individuals and development was followed microscopically. Embryos were snapfrozen in liquid nitrogen at the appropriate stage and kept at -80°C. Samples at stage 0, 8, 12, 15, 21, 26 were collected from the same fertilzation, while sample from stage 11 was independently collected from a different individual. Total RNA was prepared with the RNA II kit of Magerey Nagel and the quality was checked with the Bioanalyzer, all samples had a RIN value of higher than 9. Strand specific libraries were constructed and the sequencing (50PE) was performed with Illumina HiSeq 2000 technology (BGI, Hong Kong, China). Reads were aligned onto C.robusta genome and considered to reflect the expression of the C.robusta orthologous genes.

Conversion to Aniseed format of data from Yutaka Satou's gene expression profiles of transcription factors and signalling molecule ISH screen (Development. 2004 131:4047-58).

Original annotation :
Gene name :isl.
Expression pattern : No signal was detected

Conversion to Aniseed format of data from Yutaka Satou's gene expression profiles of transcription factors and signalling molecule ISH screen (Development. 2004 131:4047-58).

Original annotation :
Gene name :isl.
Expression pattern : No distinct zygotic signal was detected

Conversion to Aniseed format of data from Yutaka Satou's gene expression profiles of transcription factors and signalling molecule ISH screen (Development. 2004 131:4047-58).

Original annotation :
Gene name :isl.
Expression pattern : No distinct zygotic signal was detected

Conversion to Aniseed format of data from Yutaka Satou's gene expression profiles of transcription factors and signalling molecule ISH screen (Development. 2004 131:4047-58).

Original annotation :
Gene name :isl.
Expression pattern : No distinct zygotic signal was detected

Conversion to Aniseed format of data from Yutaka Satou's gene expression profiles of transcription factors and signalling molecule ISH screen (Development. 2004 131:4047-58).

Original annotation :
Gene name :isl.
Expression pattern : No distinct zygotic signal was detected

Conversion to Aniseed format of data from Yutaka Satou's gene expression profiles of transcription factors and signalling molecule ISH screen (Development. 2004 131:4047-58).

Original annotation :
Gene name :isl.
Expression pattern : No distinct zygotic signal was detected

Conversion to Aniseed format of data from Yutaka Satou's gene expression profiles of transcription factors and signalling molecule ISH screen (Development. 2004 131:4047-58).

Original annotation :
Gene name :isl.
Expression pattern : nervous system

Conversion to Aniseed format of Ghost database entry from Satou Y et al., Development 2001; 128:2893-2904. Original annotation: the entire body was stained, but the mesenchyme was relatively strongly stained.. Expression pattern: Mesenchyme.

Aniseed annotation :
This annotation conflicts with later screens, suggesting that it may be inacurate.

Conversion to Aniseed format of Ghost database entry from Satou Y et al., Development 2001; 128:2893-2904. Original annotation: the entire body was stained, but the mesenchyme was relatively strongly stained.. Expression pattern: not clear.

Aniseed annotation :
This expression pattern conflicts with later ones and may be inacurate.

Early tailbud stage wild type embryo probed for Islet expression. No picture available in the article.

Original Annotation :
Islet was expressed in the visceral ganglion (A10.57 cell pair).

Conversion to Aniseed format of data from Yutaka Satous gene expression profiles of transcription factors and signalling molecule ISH screen (Development. 2004 131:4047-58).

Original annotation :
Gene name :isl.
Expression pattern : nervous system, nerve cord.
Comment : Epidermal neurons, neurons in the palp, and neurons near the ganglion.

Aniseed annotation :
The anterior notochord cells appear stained. Not clear which epidermal neurons are stained.

At the early tailbud stage, islet is expressed in the palps.

Conversion to Aniseed format of Ghost database entry from Satou Y et al., Development 2001; 128:2893-2904. Original annotation: the entire body was stained, but the mesenchyme was relatively strongly stained.. Expression pattern: not clear.

Aniseed annotation :
Expression patterns is different from several other determinations and may be inacurate.

Dorsal view of a mid tailbud stage wild type embryo probed for Islet expression.
Red and yellow arrowheads indicate boundaries between the neck and the visceral ganglion, and between the visceral ganglion and the caudal nerve cord, respectively.

Original Annotation :
Islet was expressed in the visceral ganglion and in the tail nerve cord (A10.57 cell pair).

Conversion to Aniseed format of data from Yutaka Satous gene expression profiles of transcription factors and signalling molecule ISH screen (Development. 2004 131:4047-58).

Original annotation :
Gene name :isl.
Expression pattern : nervous system, nerve cord, notochord.
Comment : Epidermal neurons, neurons in the palp, and neurons near the ganglion

Mid-tailbud embryo probed for Islet expression. Islet is expressed in the palps, in the visceral ganglion and in the notochord.
Original annotation : Islet FISH of mid-tailbud stage embryos. They are control embryos incubated with DMSO. Nuclei are stained with Hoechst 33342.

Mid tailbud embryo where Islet>mCherry reporter (Red) was co-expressed with FoxC>YFP-caax (Green)and FoxC>lacZ. Islet enhancer is active in the palps and in the notochord.

Posterior region of the tail of a mid tailbud-stage Ciona robusta embryo, coelectroporated with Islet>unc-76::GFP (green) and Ngn>mCherry (red) reporter constructs. The second image is the un-annotated HD picture.
Original annotation: Neurogenin (Ngn) cis-regulatory sequence drives mCherry expression in A- and b- neural plate border cells, which gives rise to ependymal cells (open arrowheads) and bipolar neurons (solid arrowheads) of the posterior tail nerve cord, as well as secondary tail muscle cells (“m”). The Islet driver only drives expression in bipolar tail neurons.

Posterior region of the tail of a mid tailbud-stage Ciona robusta embryo, coelectroporated with Islet>unc-76::GFP (green) and Ngn>mCherry (red) reporter constructs. The second image is the un-annotated HD picture.
Original annotation: Neurogenin (Ngn) cis-regulatory sequence drives mCherry expression in A- and b- neural plate border cells, which gives rise to ependymal cells (open arrowheads) and bipolar neurons (solid arrowheads) of the posterior tail nerve cord, as well as secondary tail muscle cells (“m”). The Islet driver only drives expression in bipolar tail neurons.

Conversion to Aniseed format of Ghost database entry from Satou Y et al., Development 2001; 128:2893-2904. Original annotation: The trunk region was weakly stained.. Expression pattern: not clear.

Aniseed annotation :
Expression patterns is different from several other determinations and may be inacurate.

Lateral view of a WT Ciona robusta late tailbud embryo probed for Islet (KH.L152.2) expression. All embryos counterstained by fluorescent antibody stain for X-gal (red nuclei), denoting A9.30 lineage labeled by electroporation with Fgf8/17/18>lacZ.
C' is a higher magnification of C.
Original annotation: In situ hybridization of Islet in green, showing expression in A10.64 (motor neuron 2 precursor, arrowhead in C). Strong signal also seen in other tissues, including notochord (broken outline in C), palps, pharyngeal mesoderm and bipolar tail neurons.

Lateral view of the visceral ganglion of a WT Ciona robusta late tailbud stage embryo probed for Islet expression (green) and electroporated with Fgf8/17/18>nlsLacZ (labels in red nuclei from A9.30 lineage) and COE>mCherry (no staining, control experiment).
Original annotation: Islet is expressed in A10.64 (motor neuron 2 precursor, open arrow) and also in the notochord, just ventral to the nerve cord.

Lateral view of the visceral ganglion of a WT Ciona robusta late tailbud embryo co-electroporated with Islet>unc76::GFP (green) and Fgf8/17/18>unc76::mCherry (red) reporter constructs.
Original annotation:A9.30 lineage is labeled by electroporation of Fgf8/17/18>mCherry.
The upstream enhancer of Islet drives reporter gene expression in A10.64 (motor neuron 2 precursor).

Lateral view of the visceral ganglion of a WT Ciona robusta late tailbud embryo co-electroporated with Islet>unc76::GFP (green) and Fgf8/17/18>unc76::mCherry (red) reporter constructs.
Original annotation: A9.30 lineage is labeled by electroporation of Fgf8/17/18>mCherry.
The upstream enhancer of Islet drives reporter gene expression in A10.64 (motor neuron 2 precursor).

WT late tailbud embryo probed for Emx expression (green), and islet expression (red).
Original annotation : Double ISH of late tailbud embryo. Islet probe (DIG-labeled, red) is detected in spots corresponding to the presumptive palp protrusions and is flanked by rings of Emx expression (fluorescein-labeled, green).
(D')Close-up view focused on palp region. Nuclei are stained with Hoechst 33342. (D'', D''') in situ signal of respective single probes shown for clarity. Scale bars: 10 μm.

WT late tailbud embryo probed for islet expression (red).
Original annotation : In situ hybridization reveals Islet transcript in the three presumptive palps at late tailbud stage. Image shown is a confocal z-projection of 15 slices.
Two of the three presumptive palps, indicated by single and double arrows, are shown in magnified view in panels B-C′. (B-C′) Confocal z-projections of two slices focused on an individual palp protrusion. (B,C) Hoechst 33342 counterstaining allows visualization of protruding palp. (B′,C′) Same images as in B and C, but with Islet in situ signal overlaid. (D-E′) Islet>mCherry reporter is expressed in the palps of late tail bud embryo. Image shown is a confocal z-projection of four slices focused on one palp. Phalloidin staining reveals cell cortices. Cells expressing Islet reporter are elongated in comparison to neighboring non- expressing cells.

Ventral view of the head of a WT Ciona robusta late tailbud embryo probed for Islet expression (green) and transfected with Mesp>LacZ (red) to visualize the B7.5 lineage.
Islet is expressed in several tissues including atrial siphon muscles (ASM) precursors, trunk endoderm (asterisks), oral siphon and palps.
Solid arrowheads point to the lateral trunk ventral cells (TVCs) (i.e. the ASM precursors) and open arrowheads point to the median TVCs (i.e. the heart precursors).

mCherry expression driven by islet cis-regulatory sequence in a late tailbud embryo.
Original annotation : Islet>mCherry reporter is expressed in the palps of late tailbud embryo. Image shown is a confocal z-projection of four slices focused on one palp. Phalloidin staining reveals cell cortices. Cells expressing Islet reporter are elongated in comparison to neighboring non-expressing cells. Scale bar: 10 μm

Lateral view of the visceral ganglion of a WT Ciona robusta late tailbud embryo co-electroporated with Onecut>unc76::GFP (green) and Islet>unc76::mCherry (red) reporter constructs.
Onecut enhancer drives GFP expression in differentiating motor neurons and nerve cord cells.
Original annotation:A −4.2 kb fragment immediately upstream of the transcription start site of the neuronal differentiation factor Onecut (also known as HNF6) drives reporter gene expression in differentiating neurons and nerve cord cells. This labels frondose endplates that contact the sides of the dorsal and anterior tail muscle cells.
Figure:
(B) Co-electroporation with Islet>mCherry reveals that these endplates do not belong to A10.64, which is strongly stained by this construct. (C) Magnification of dotted area in A. (C′) Magnification of dotted area in B. (C′′) Merged image of C and C′, showing that A10.64 might even be contacting and synapsing onto the frondose endplates (arrow).

Lateral view of the visceral ganglion of a WT Ciona robusta late tailbud embryo co-electroporated with Onecut>unc76::GFP (green) and Islet>unc76::mCherry (red) reporter constructs.
Islet enhancer drives mCherry expression in A10.64 differentiating motor neuron 2.
Original annotation of the figure: (A) A −4.2 kb fragment immediately upstream of the transcription start site of the neuronal differentiation factor Onecut (also known as HNF6) drives reporter gene expression in differentiating neurons and nerve cord cells. This labels frondose endplates that contact the sides of the dorsal and anterior tail muscle cells.
(B) Co-electroporation with Islet>mCherry reveals that these endplates do not belong to A10.64, which is strongly stained by this construct. (C) Magnification of dotted area in A.
(C′) Magnification of dotted area in B.
(C′′) Merged image of C and C′, showing that A10.64 might even be contacting and synapsing onto the frondose endplates (arrow).

Ventral view of the head of a WT Ciona robusta larva probed for Islet expression (green) and transfected with Mesp>LacZ (red) to visualize the B7.5 lineage.
Islet expression persists in the oral siphon, palps (arrows), parts of the trunk endoderm (asterisks), and ASM precusors (solid arrowheads), but is not detected above background levels in the heart precursors (open arrowheads).

Hatching larva electroporated with FoxC>H2B:Cherry (Red) and Islet>YFPcaax (Green) reporter plasmids and stained with phalloidin in control conditions (ie co-electroporated with FoxC>LacZ).
Original annotation : Control embryo expressing FoxC>lacZ shows Islet reporter expression in protrusive palps (YFP-caax is a membrane-targeted YFP).

Lateral view of A10.64 (motor neuron 2) from the visceral ganglion of a WT Ciona robusta swimming tadpole electroporated with Islet>unc76::mCherry.
Original annotation:A10.64, as revealed by Islet>mCherry and in accordance with previous studies, displays a cell body more elongated along its anterior-posterior axis than the other VG neurons (Imai and Meinertzhagen, 2007; Okada et al., 2002), but does not form prominent endplates.

Lateral view of the visceral ganglion of a WT Ciona robusta larva co-electroporated with Dmbx>unc76::CFP (blue), Vsx>unc76::mCherry (red), Nk6>unc76::YFP (green), and Islet>unc76::mCherry and Islet>unc76::CFP (purple) reporter constructs.
Dmbx enhancer drives CFP expression in A12.239 motor neuron.
Original annotation:Five motor neurons are simultaneously visualized and distinguishable from one another. Green channel was imaged on a different focal plane than blue or red, to visualize Nk6>YFP-labeled endplates. Yellow color only indicates overlap of cell bodies, not colocalization.
Diagram of the Ciona visceral ganglion ‘brainbow’ showing the neurons from the picture and their unique morphological traits, such as contralateral projection of A12.239 pair, smaller cell body of A13.474, frondose endplates of A11.118, dendrites on A11.117, and elongated cell body and smaller endplates of A10.64 (motor neuron 2). Pale-green shading indicates left side of the larva. Pink fibers represent tail muscle.

Lateral view of the visceral ganglion of a WT Ciona robusta larva co-electroporated with Dmbx>unc76::CFP (blue), Vsx>unc76::mCherry (red), Nk6>unc76::YFP (green), and Islet>unc76::mCherry and Islet>unc76::CFP (purple) reporter constructs.
Vsx enhancer drives mCherry expression in A13.474 and A11.117 motor neurons.
Original annotation:Five motor neurons are simultaneously visualized and distinguishable from one another. Green channel was imaged on a different focal plane than blue or red, to visualize Nk6>YFP-labeled endplates. Yellow color only indicates overlap of cell bodies, not colocalization.
(B) Diagram showing the neurons from the picture and their unique morphological traits, such as contralateral projection of A12.239 pair, smaller cell body of A13.474, frondose endplates of A11.118, dendrites on A11.117, and elongated cell body and smaller endplates of A10.64 (motor neuron 2). Pale-green shading indicates left side of the larva. Pink fibers represent tail muscle.

Lateral view of the visceral ganglion of a WT Ciona robusta larva co-electroporated with Dmbx>unc76::CFP (blue), Vsx>unc76::mCherry (red), Nk6>unc76::YFP (green), and Islet>unc76::mCherry and Islet>unc76::CFP (purple) reporter constructs.
Nk6 enhancer drives YFP expression in A11.118 motor neuron (green).
Original annotation:Five motor neurons are simultaneously visualized and distinguishable from one another. Green channel was imaged on a different focal plane than blue or red, to visualize Nk6>YFP-labeled endplates. Yellow color only indicates overlap of cell bodies, not colocalization.
Diagram of the Ciona visceral ganglion ‘brainbow’ showing the neurons from the picture and their unique morphological traits, such as contralateral projection of A12.239 pair, smaller cell body of A13.474, frondose endplates of A11.118, dendrites on A11.117, and elongated cell body and smaller endplates of A10.64 (motor neuron 2). Pale-green shading indicates left side of the larva. Pink fibers represent tail muscle.

Lateral view of the visceral ganglion of a WT Ciona robusta larva co-electroporated with Dmbx>unc76::CFP (blue), Vsx>unc76::mCherry (red), Nk6>unc76::YFP (green), and Islet>unc76::mCherry and Islet>unc76::CFP (purple) reporter constructs.
Islet enhancer drives mCherry and CFP expression in A10.64 motor neuron 2 (purple).
Original annotation:Five motor neurons are simultaneously visualized and distinguishable from one another. Green channel was imaged on a different focal plane than blue or red, to visualize Nk6>YFP-labeled endplates. Yellow color only indicates overlap of cell bodies, not colocalization.
Diagram of the Ciona visceral ganglion ‘brainbow’ showing the neurons from the picture and their unique morphological traits, such as contralateral projection of A12.239 pair, smaller cell body of A13.474, frondose endplates of A11.118, dendrites on A11.117, and elongated cell body and smaller endplates of A10.64. Pale-green shading indicates left side of the larva. Pink fibers represent tail muscle.

Lateral view of the visceral ganglion of a WT Ciona robusta larva co-electroporated with Dmbx>unc76::CFP (blue), Vsx>unc76::mCherry (red), Nk6>unc76::YFP (green), and Islet>unc76::mCherry and Islet>unc76::CFP (purple) reporter constructs.
Islet enhancer drives mCherry and CFP expression in A10.64 motor neuron 2 (purple).
Original annotation:Five motor neurons are simultaneously visualized and distinguishable from one another. Green channel was imaged on a different focal plane than blue or red, to visualize Nk6>YFP-labeled endplates. Yellow color only indicates overlap of cell bodies, not colocalization.
Diagram of the Ciona visceral ganglion ‘brainbow’ showing the neurons from the picture and their unique morphological traits, such as contralateral projection of A12.239 pair, smaller cell body of A13.474, frondose endplates of A11.118, dendrites on A11.117, and elongated cell body and smaller endplates of A10.64. Pale-green shading indicates left side of the larva. Pink fibers represent tail muscle.

Lateral view of the visceral ganglion of a WT Ciona robusta larva co-electroporated with Dmbx>unc76::CFP (blue), Vsx>unc76::mCherry (red), Nk6>unc76::YFP (green), and Islet>unc76::mCherry and Islet>unc76::CFP (purple) reporter constructs.
Dmbx enhancer drives CFP expression in A12.239 motor neuron.
Original annotation:Five motor neurons are simultaneously visualized and distinguishable from one another. Green channel was imaged on a different focal plane than blue or red, to visualize Nk6>YFP-labeled endplates. Yellow color only indicates overlap of cell bodies, not colocalization.
Diagram of the Ciona visceral ganglion ‘brainbow’ showing the neurons from the picture and their unique morphological traits, such as contralateral projection of A12.239 pair, smaller cell body of A13.474, frondose endplates of A11.118, dendrites on A11.117, and elongated cell body and smaller endplates of A10.64 (motor neuron 2). Pale-green shading indicates left side of the larva. Pink fibers represent tail muscle.

Lateral view of the visceral ganglion of a WT Ciona robusta larva co-electroporated with Dmbx>unc76::CFP (blue), Vsx>unc76::mCherry (red), Nk6>unc76::YFP (green), and Islet>unc76::mCherry and Islet>unc76::CFP (purple) reporter constructs.
Vsx enhancer drives mCherry expression in A13.474 and A11.117 motor neurons.
Original annotation:Five motor neurons are simultaneously visualized and distinguishable from one another. Green channel was imaged on a different focal plane than blue or red, to visualize Nk6>YFP-labeled endplates. Yellow color only indicates overlap of cell bodies, not colocalization.
(B) Diagram showing the neurons from the picture and their unique morphological traits, such as contralateral projection of A12.239 pair, smaller cell body of A13.474, frondose endplates of A11.118, dendrites on A11.117, and elongated cell body and smaller endplates of A10.64 (motor neuron 2). Pale-green shading indicates left side of the larva. Pink fibers represent tail muscle.

Lateral view of the visceral ganglion of a WT Ciona robusta larva co-electroporated with Dmbx>unc76::CFP (blue), Vsx>unc76::mCherry (red), Nk6>unc76::YFP (green), and Islet>unc76::mCherry and Islet>unc76::CFP (purple) reporter constructs.
Nk6 enhancer drives YFP expression in A11.118 motor neuron (green).
Original annotation:Five motor neurons are simultaneously visualized and distinguishable from one another. Green channel was imaged on a different focal plane than blue or red, to visualize Nk6>YFP-labeled endplates. Yellow color only indicates overlap of cell bodies, not colocalization.
Diagram of the Ciona visceral ganglion ‘brainbow’ showing the neurons from the picture and their unique morphological traits, such as contralateral projection of A12.239 pair, smaller cell body of A13.474, frondose endplates of A11.118, dendrites on A11.117, and elongated cell body and smaller endplates of A10.64 (motor neuron 2). Pale-green shading indicates left side of the larva. Pink fibers represent tail muscle.

Lateral view of the visceral ganglion of a WT Ciona robusta larva co-electroporated with Dmbx>unc76::CFP (blue), Vsx>unc76::mCherry (red), Nk6>unc76::YFP (green), and Islet>unc76::mCherry and Islet>unc76::CFP (purple) reporter constructs.
Islet enhancer drives mCherry and CFP expression in A10.64 motor neuron 2 (purple).
Original annotation:Five motor neurons are simultaneously visualized and distinguishable from one another. Green channel was imaged on a different focal plane than blue or red, to visualize Nk6>YFP-labeled endplates. Yellow color only indicates overlap of cell bodies, not colocalization.
Diagram of the Ciona visceral ganglion ‘brainbow’ showing the neurons from the picture and their unique morphological traits, such as contralateral projection of A12.239 pair, smaller cell body of A13.474, frondose endplates of A11.118, dendrites on A11.117, and elongated cell body and smaller endplates of A10.64. Pale-green shading indicates left side of the larva. Pink fibers represent tail muscle.

Lateral view of the visceral ganglion of a WT Ciona robusta larva co-electroporated with Dmbx>unc76::CFP (blue), Vsx>unc76::mCherry (red), Nk6>unc76::YFP (green), and Islet>unc76::mCherry and Islet>unc76::CFP (purple) reporter constructs.
Islet enhancer drives mCherry and CFP expression in A10.64 motor neuron 2 (purple).
Original annotation:Five motor neurons are simultaneously visualized and distinguishable from one another. Green channel was imaged on a different focal plane than blue or red, to visualize Nk6>YFP-labeled endplates. Yellow color only indicates overlap of cell bodies, not colocalization.
Diagram of the Ciona visceral ganglion ‘brainbow’ showing the neurons from the picture and their unique morphological traits, such as contralateral projection of A12.239 pair, smaller cell body of A13.474, frondose endplates of A11.118, dendrites on A11.117, and elongated cell body and smaller endplates of A10.64. Pale-green shading indicates left side of the larva. Pink fibers represent tail muscle.

Dorsal view of electroporated Ciona robusta larva exhibiting mosaic incorporation (left side) of Islet>GFP (green) and MesP>H2B∷mCherry (red) transgenes at 20 hpf, before the migration of the lateral TVCs.
Mesp enhancer is active in anterior tail muscles (asterisk) and in atrial siphon muscles precursors. Islet>GFP expression is restricted to lateral TVCs. Dotted line indicates midline; M and L indicate medial and lateral, respectively.

Dorsal view of electroporated Ciona robusta larva exhibiting mosaic incorporation (left side) of Islet>GFP (green) and MesP>H2B∷mCherry (red) transgenes at 20 hpf, before the migration of the lateral TVCs.
Mesp enhancer is active in anterior tail muscles (asterisk) and in atrial siphon muscles precursors. Islet>GFP expression is restricted to lateral TVCs. Dotted line indicates midline; M and L indicate medial and lateral, respectively.

Ciona robusta larva electroporated with Islet -7216/-3950::bpFOG>unc76::GFP, showing expression in atrial siphon muscle precursors (arrowhead) and cholinergic neurons (arrows).
This Islet enhancer is in green in the diagram.

Ciona robusta larva electroporated with Islet -5915/-5396::bpFOG>unc76::GFP, showing expression in bipolar tail neurons (BTNs) only.
This Islet enhancer is in purple in the diagram.

No embryo picture in the article.
The Islet -7216/+1::bpFOG>unc76::GFP reporter construct drives strong GFP expression in atrial siphon muscle precursors (ASM), notochord (Noto), bipolar tail neurons (BTN) and cholinergic neurons (ChN) of the central nervous system of Ciona robusta larvae. This Islet enhancer is in red in the diagram.
Original legend: Expression is given as percentage of transfected larvae with visible GFP fluorescence in TVCs at 24 hpf (n = 35 to 195). Minus sign (“-“) indicates no or very little (<4%) expression. Single plus sign (“+”) indicates expression in 10 to 50% of transfected larvae. Double plus sign (“++”) indicates expression in greater than 50% of transfected larvae. Plus signs without percentage value indicate this expression was visually estimated to be >75% of transfected larvae.

No embryo picture in the article.
The Islet -3950/+1::bpFOG>unc76::GFP reporter construct drives strong GFP expression in the notochord (Noto) of Ciona robusta larvae. This Islet enhancer is in yellow in the diagram.
Original legend: Expression is given as percentage of transfected larvae with visible GFP fluorescence in TVCs at 24 hpf (n = 35 to 195). Minus sign (“-“) indicates no or very little (<4%) expression. Single plus sign (“+”) indicates expression in 10 to 50% of transfected larvae. Double plus sign (“++”) indicates expression in greater than 50% of transfected larvae. Plus signs without percentage value indicate this expression was visually estimated to be >75% of transfected larvae.

No embryo picture in the article.
The Islet -5035/-3950::bpFOG>unc76::GFP reporter construct drives GFP expression in atrial siphon muscle precursors (ASM) and cholinergic neurons (ChN) of Ciona robusta larvae. This Islet enhancer is in blue in the diagram.
Original legend: Expression is given as percentage of transfected larvae with visible GFP fluorescence in TVCs at 24 hpf (n = 35 to 195). Minus sign (“-“) indicates no or very little (<4%) expression. Single plus sign (“+”) indicates expression in 10 to 50% of transfected larvae. Double plus sign (“++”) indicates expression in greater than 50% of transfected larvae. Plus signs without percentage value indicate this expression was visually estimated to be >75% of transfected larvae.

No embryo picture in the article.
The Islet -6240/-5035::bpFOG>unc76::GFP reporter construct drives strong GFP expression in atrial siphon muscle precursors (ASM) and bipolar tail neurons of Ciona robusta larvae. This Islet enhancer is in grey in the diagram.
Original legend: Expression is given as percentage of transfected larvae with visible GFP fluorescence in TVCs at 24 hpf (n = 35 to 195). Minus sign (“-“) indicates no or very little (<4%) expression. Single plus sign (“+”) indicates expression in 10 to 50% of transfected larvae. Double plus sign (“++”) indicates expression in greater than 50% of transfected larvae. Plus signs without percentage value indicate this expression was visually estimated to be >75% of transfected larvae.

No embryo picture in the article.
The Islet -5396/-4982::bpFOG>unc76::GFP reporter construct drives GFP expression in atrial siphon muscle precursors (ASM) of Ciona robusta larvae. This Islet enhancer is in orange in the diagram.
Original legend: Expression is given as percentage of transfected larvae with visible GFP fluorescence in TVCs at 24 hpf (n = 35 to 195). Minus sign (“-“) indicates no or very little (<4%) expression. Single plus sign (“+”) indicates expression in 10 to 50% of transfected larvae. Double plus sign (“++”) indicates expression in greater than 50% of transfected larvae. Plus signs without percentage value indicate this expression was visually estimated to be >75% of transfected larvae.

No embryo picture in the article.
The Islet -7216/-6240::bpFOG>unc76::GFP reporter construct drives no GFP expression Ciona robusta larvae. This Islet enhancer is in cyan in the diagram.
Original legend: Expression is given as percentage of transfected larvae with visible GFP fluorescence in TVCs at 24 hpf (n = 35 to 195). Minus sign (“-“) indicates no or very little (<4%) expression. Single plus sign (“+”) indicates expression in 10 to 50% of transfected larvae. Double plus sign (“++”) indicates expression in greater than 50% of transfected larvae. Plus signs without percentage value indicate this expression was visually estimated to be >75% of transfected larvae.

Lateral view of the head of a Ciona robusta larva transfected with MesP>H2B::mCherry (red nuclei) and Islect>GFP (green) reporter constructs, and FoxF[TVC]>LacZ as a control.
Mesp enhancer is active in B7.5 lineage, [ie. heart primordium (arrowhead) and atrial siphon muscle (ASM) precursors (arrow)], and Islet enhancer in atrial ASM precursors and in the visceral ganglion.

Lateral view of the head of a Ciona robusta larva transfected with MesP>H2B::mCherry (red nuclei) and Islect>GFP (green) reporter constructs, and FoxF[TVC]>LacZ as a control.
Mesp enhancer is active in B7.5 lineage, [ie. heart primordium (arrowhead) and atrial siphon muscle (ASM) precursors (arrow)], and Islet enhancer in atrial ASM precursors and in the visceral ganglion.

Lateral view of a WT Ciona robusta larva electroporated with islet_BTN>unc-76::eGFP.
GFP is fused to UNC-76 tags to ensure even labelling of axons. GFP signalling is observed in bipolar tail neurons (BTNs) of Ciona larvae.
Original annotation : Larva with a BTN labelled by Islet BTN>unc-76::eGFP (green). BTNs form axon fascicles that extend along the length of the tail on either side of the neural tube. Bottom, enlarged view of BTN above. Scale bars, 75 μm (top); 25 μm (bottom).

Ventral view of the head of a WT Ciona robusta larva probed for Islet expression (green) and transfected with Mesp>LacZ (red) to visualize the B7.5 lineage.
Islet expression persists in the oral siphon, palps (arrows), parts of the trunk endoderm (asterisks), and ASM precusors (solid arrowheads), but is not detected above background levels in the heart precursors (open arrowheads).

mCherry expression driven by islet cis-regulatory sequence in mature larva.
Original annotation : Islet>mCherry reporter expression in a mature larva, with phalloidin counterstaining. G shows phalloidin only; arrow indicates digitiform protrusions at tips of palps. G′ shows Islet reporter only,revealing that the digitiform protrusions (arrow) derive from the Islet+ cells.
Aniseed annotation : there is no annotation for this entry because the corresponding anatomical ontology for this stage is not yet available in ANISEED.

Lateral view of a Ciona robusta larva electroporated with Islet>GFP (green) and MesP>H2B∷mCherry (red) transgenes at 24hpf, after migration of Islet>GFP-positive lateral TVC descendants around the atrial siphon primordium.
Mesp enhancer is active in anterior tail muscles (asterisk) and in atrial siphon muscles precursors. Islet>GFP expression is restricted to lateral TVCs. D and V indicate dorsal and ventral, respectively.

Lateral view of a Ciona robusta larva electroporated with Islet>GFP (green) and MesP>H2B∷mCherry (red) transgenes at 24hpf, after migration of Islet>GFP-positive lateral TVC descendants around the atrial siphon primordium.
Mesp enhancer is active in anterior tail muscles (asterisk) and in atrial siphon muscles precursors. Islet>GFP expression is restricted to lateral TVCs. D and V indicate dorsal and ventral, respectively.

Ciona robusta juvenile (~100 hpf) raised from embryo transfected with Islet>H2B∷mCherry (red), with transgene expression visible around atrial siphons (arrow) and longitudinal muscles (arrowheads), but not heart (Ht). F-actin is stained by phallacidin (blue-green). Scale bar, 100 μm.

Ciona robusta juvenile (~100 hpf) raised from embryo transfected with Islet>H2B∷mCherry (B, red) and Mesp>LacZ (A, red).C is the merge image.
H2B::mCherry is observed in atrial siphon muscles and longitudinal muscles, but not heart.
Differentiated B7.5 descendants (heart, atrial siphon muscles, and longitudinal muscles) are visualized using Mesp>LacZ staining (red).
Original comment: . Insets c’ and c’’ are magnified views of corresponding dashed line boxes. Note co-localization of βgal immunostain and H2B::mCherry around atrial siphons (c’) but not in the heart (c’’). Asterisks denote βgal+, H2B::mCherry+ nuclei of longitudinal muscles in which βgal was obscured in the merged image. Arrow in c’’ show Islet+ cells of unknown origin in the endostyle. Islet>H2B::mCherry is also seen in the re-absorbed tail (t). Juvenile was counterstained by phallacidin:BODIPY FL conjugate.

Ciona robusta juvenile (~100 hpf) raised from embryo transfected with Islet>H2B∷mCherry (B, red) and Mesp>LacZ (A, red).C is the merge image.
H2B::mCherry is observed in atrial siphon muscles and longitudinal muscles, but not heart.
Differentiated B7.5 descendants (heart, atrial siphon muscles, and longitudinal muscles) are visualized using Mesp>LacZ staining (red).
Original comment: Insets c’ and c’’ are magnified views of corresponding dashed line boxes. Note co-localization of βgal immunostain and H2B::mCherry around atrial siphons (c’) but not in the heart (c’’). Asterisks denote βgal+, H2B::mCherry+ nuclei of longitudinal muscles in which βgal was obscured in the merged image. Arrow in c’’ show Islet+ cells of unknown origin in the endostyle. Islet>H2B::mCherry is also seen in the re-absorbed tail (t). Juvenile was counterstained by phallacidin:BODIPY FL conjugate.

Conversion to Aniseed format of Ghost database entry from Ogasawara et al., Dev Genes Evol. 2002; 212:173-185. Original annotation: . Expression pattern: Epidermis