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 :EphrinA-d.
Expression pattern : Unlocalized signal was detected.

Aniseed annotation :
Signal is barely detectable on picture.

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 :EphrinA-d.
Expression pattern : No distinct zygotic signal was detected

Scale bar, 100 μm

Vegetal and animal view of a 16-cell stage embryo stained with EphrinA-d expression. Expression of EphrinA-d is observed in the a5.3, a5.4, b5.3 and b5.4 cell pairs.

Animal view of a WT Ciona robusta 16-cell stage embryo probed for Efna.d expression.
Efna.d is expressed in a5.3, a5.4, A6.2 and A6.4 blastomers.

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 :EphrinA-d.
Expression pattern : a5.3, a5.4, b5.3, b5.4

Lateral (anterior to the left) and animal (anterior up) view of a 16 cell stage embryo probed for Ci-ephrin-Ad expression.

Animal view of a 16-cell stage embryo showing expression of Ci-ephrin-Ad.

Animal and vegetal views of a 16-cell stage embryo probed for Efna.d expression.
Original annotation: In normal embryos and embryos injected with the control MO, Efna.d are expressed in the AD.

Illustrations on the left depict the constructs. The numbers indicate the relative nucleotide positions from the transcription start site of Efna.d. Mutant Tcf7-binding sites are indicated by X. Graphs show the percentage of blastomeres expressing the reporter in the vegetal blastomeres and in the animal blastomeres. Note that not all cells could express the reporter because of mosaic incorporation of the electroporated plasmid, and not all embryos could express the reporter either.

First picture
Illustrations on the left depict the constructs. The numbers indicate the relative nucleotide positions from the transcription start site of Efna.d. Mutant Tcf7-binding sites are indicated by X. Graphs show the percentage of blastomeres expressing the reporter in the vegetal blastomeres and in the animal blastomeres. Note that not all cells could express the reporter because of mosaic incorporation of the electroporated plasmid, and not all embryos could express the reporter either.
Second picture
Images showing Gfp expression in embryos electroporated with the construct shown in the first picture. Scale bar, 100 μm.
"A series of deletion constructs of the upstream sequence of Efna.d revealed that the region between −419 and −220 was essential for driving reporter expression in the AD."

Illustrations on the left depict the constructs. The numbers indicate the relative nucleotide positions from the transcription start site of Efna.d. Mutant Tcf7-binding sites are indicated by X. Graphs show the percentage of blastomeres expressing the reporter in the vegetal blastomeres and in the animal blastomeres. Note that not all cells could express the reporter because of mosaic incorporation of the electroporated plasmid, and not all embryos could express the reporter either.

Illustrations on the left depict the constructs. The numbers indicate the relative nucleotide positions from the transcription start site of Efna.d. Mutant Tcf7-binding sites are indicated by X. Graphs show the percentage of blastomeres expressing the reporter in the vegetal blastomeres and in the animal blastomeres. Note that not all cells could express the reporter because of mosaic incorporation of the electroporated plasmid, and not all embryos could express the reporter either.

Illustrations on the left depict the constructs. The numbers indicate the relative nucleotide positions from the transcription start site of Efna.d. Mutant Tcf7-binding sites are indicated by X. Graphs show the percentage of blastomeres expressing the reporter in the vegetal blastomeres and in the animal blastomeres. Note that not all cells could express the reporter because of mosaic incorporation of the electroporated plasmid, and not all embryos could express the reporter either.

First picture
Illustrations on the left depict the constructs. The numbers indicate the relative nucleotide positions from the transcription start site of Efna.d. Mutant Tcf7-binding sites are indicated by X. Graphs show the percentage of blastomeres expressing the reporter in the vegetal blastomeres and in the animal blastomeres. Note that not all cells could express the reporter because of mosaic incorporation of the electroporated plasmid, and not all embryos could express the reporter either.
Second picture
Images showing Gfp expression in embryos electroporated with the construct shown in the first picture. Scale bar, 100 μm.
"A series of deletion constructs of the upstream sequence of Efna.d revealed that the region between −419 and −220 was essential for driving reporter expression in the AD."
"Mutations introduced into three or four putative Gata.a-binding sites within this region impaired the activity of this regulatory region."

Illustrations on the left depict the constructs. The numbers indicate the relative nucleotide positions from the transcription start site of Efna.d. Mutant Tcf7-binding sites are indicated by X. Graphs show the percentage of blastomeres expressing the reporter in the vegetal blastomeres and in the animal blastomeres. Note that not all cells could express the reporter because of mosaic incorporation of the electroporated plasmid, and not all embryos could express the reporter either.
"Mutations introduced into three or four putative Gata.a-binding sites within this region impaired the activity of this regulatory region."

Lateral (anterior to the left) and animal (anterior up) view of a 24 cell stage embryo probed for Ci-ephrin-Ad expression. The position of A6.2 blastomere is indicated by the red line.

The DNA fragment, containing 1648-bp upstream region, drove transcription of lacZ reporter gene in 8 cells in the animal hemisphere of the 16-cell embryo.

The DNA fragment, containing 545-bp upstream region, drove transcription of lacZ reporter gene in 8 cells in the animal hemisphere of the 16-cell embryo.

The DNA fragment, containing 497-bp upstream region, drove transcription of lacZ reporter gene in 8 cells in the animal hemisphere of the 16-cell embryo.

The DNA fragment, containing 447-bp upstream region, drove transcription of lacZ reporter gene in 8 cells in the animal hemisphere of the 16-cell embryo.

The DNA fragment, containing 397-bp upstream region, drove transcription of lacZ reporter gene in 8 cells in the animal hemisphere of the 16-cell embryo.

The DNA fragment, containing 333-bp upstream region, did not drive transcription of lacZ reporter gene in the 16-cell embryo.

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 :EphrinA-d.
Expression pattern : a6.5, a6.6, a6.7, a6.8, b6.6, b6.8

Animal view of a 32-cell stage embryo, showing expression of Ci-ephrin-Ad.

Animal view of a late-32 cell stage embryo stained with ephrin-Ad expression. Expression in the animal cells. No expression in vegetal view.

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 :EphrinA-d.
Expression pattern : No distinct zygotic signal was detected.
Comment : Some embryos expressed this gene weakly in the animal blastomeres but some embryos did not.

Animal view of a 64-cell stage embryo showing expression of Ci-ephrin-Ad. Expression diminishes at this stage.

Views of 64-cell stage embryos showing reporter expression of the region upstream of EphrinA-d in all b-line cells.

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 :EphrinA-d.
Expression pattern : No distinct zygotic signal was detected

Views of 110-cell stage embryos showing reporter expression of the region upstream of EphrinA-d in b-cells.

Vegetal view of a late gastrula stage wild type embryo probed for Ci-ephrinA-d expression.

Original Annotation :
Ci-ephrinA-d was expressed in A9.29 cell pairs.

Aniseed Annotation :
Expression was also observed in row V and row VI of the neural plate.

Original annotation :
Posterior secondary muscle lineage (A9.31 line), row V neural plate, row VI neural plate.

Aniseed annotation:
Data provided by the author, not available in the article.

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 :EphrinA-d.
Expression pattern : No distinct zygotic signal was detected.

Aniseed annotation :
Expression in the anterior neural plate can be seen on the picture.

At the late gastrula stage, EphrinA-d is expressed in rows VI and V of the neural plate.

At the late gastrula stage, EphrinA-d is expressed in cells from rows V and VI of the neural plate
Original annotation : The expression of EphrinA-d in control embryo at the late gastrula stage (shown in animal view). The expression is illustrated schematically in the lower right by red (palps) and blue (brain).

Views of late gastrula stage embryos showing reporter expression of the region upstream of EphrinA-d in ectoderm except for a9.34 and a9.38.

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 :EphrinA-d.
Expression pattern : nervous system.

Aniseed annotation :
In addition to the anterior a-line neural plate expression, a patch of expression appears in the b-line midline: is it in the neural plate, or rather in an uncharacterised b-line epidermal neuronal population?

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 :EphrinA-d.
Expression pattern : nervous system, nerve cord.
Comment : The expession was observed around the papilla, ganglion and in the posterior part of nerve cord.

Aniseed annotation :
Expression is apparent in the palp areas, the anterior sensory vesicle, part of the visceral ganglion and part of the posterior tail dorsal midline epidermis (neuronal population?).

Dorsal view of an early tailbud stage wild type embryo probed for Ci-ephrinA-d 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 :
Ci-ephrinA-d was expressed in A11.126-8 (lateral nerve cord) and in A11.118 cell pairs (visceral ganglion) .

Views of early tailbud stage embryos showing reporter expression of the region upstream of EphrinA-d in ectodermal cells and mesenchyme.

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 :EphrinA-d.
Expression pattern : nervous system, nerve cord.
Comment : The expession was observed around the papilla, ganglion and in the posterior part of nerve cord.

Mid tailbud stage wild type embryo probed for Ci-ephrinA-d expression. No picture available in the article.

Original Annotation :
Ci-ephrinA-d expression in the visceral ganglion was observed in A12.240 and A11.118 cell pairs.

Original annotation:
Expression in the tail nerve cord is restricted to the posterior two thirds.

Aniseed annotation:
Data provided by the author, not available in the article.

Views of larvae showing reporter expression of the region upstream of EphrinA-d in ectodermal cells and mesenchyme.

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