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 :Snail.
Expression pattern : No signal was detected

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 :Snail.
Expression pattern : No signal was detected

Vegetal view of a WT Ciona robusta 16-cell stage embryo probed for Snail expression.
Snail is not yet expressed at this stage (no expression observed in the 69 observed embryos).
Scale bar: 100 μm

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 :Snail.
Expression pattern : No distinct zygotic signal was detected

Vegetal (left) and posterior (right) views of a WT Ciona robusta 32-cell stage embryo probed for Snail expression.
Snail is expressed in B6.1, B6.2 and B6.4 blastomer pairs.
Scale bar: 100 μm

Vegetal view of a 32-cell stage wild type embryo stained for Ci-Sna mRNA by in situ hybridization.

Conversion to Aniseed format of Ghost database entry from Fujiwara S et al., Mech Dev. 2002; 112Gene name: 115-127. Original annotation: . Expression pattern: B6.2 B6.4

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 :Snail.
Expression pattern : B6.1, B6.2, B6.4

Vegetal view of a late 32-cell stage embryo. The transcript of ci-snail was detected from the 32-cell stage onwards in B6.1, B6.2 and B6.4 blastomeres.

Vegetal view of a WT Ciona robusta late 32-cell stage embryo probed for Snail expression.
Expression is observed in B6.1, B6.2 and B6.4 blastomere pairs.
In schematic representation of gene expression, notochord and nerve cord precursors mother cells are in purple. Blastomeres with gene expression are marked by dots.

Vegetal view of a late 32-cell stage embryo stained for Ci-Sna mRNA by in situ hybridization.

Vegetal view of a 32-cell stage embryo, electroporated with a lacZ transgene containing the 3,2kb of the 5' flanking Ci-Sna promoter sequence, with a small internal deletion within the full length promoter wich removes the 262bp-enhancer (-618 -356) and assayed by ISH against lacZ. There is a severe loss of activity compared with wild-type Ci-Sna -3.2 kb 15th codon region.

Vegetal view of a 44-cell embryo electroporated with pCi-sna262-fkh::NLS-lacZ and probed with a lacZ RNA probe. Expression is detected in the B4.1 derived muscle and mesenchyme precursors. B7.5 seems not to be labelled.

Vegetal view of a 44-cell stage embryo, electroporated with a lacZ transgene containing the 3,2kb of the 5' flanking Ci-Sna promoter sequence. Intense staining is detected in the progenitors of the tail muscles and mesenchyme.

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 :Snail.
Expression pattern : B7.1, B7.2, B7.3, B7.4, B7.7, B7.8.

Aniseed annotation : We can barely see B7.1 but we can see B7.4, B7.7, B7.8 in the picture.

Vegetal view of a WT Ciona robusta 64-cell stage embryo probed for Snail expression.
Expression is observed in B7.3, B7.4, B7.7 and B7.8 blastomere pairs.
In schematic representation of gene expression, notochord precursors are shown in red. Blastomeres with gene expression are marked by dots.

Picture 1 : Vegetal view of a 64-cell stage embryo stained for Ci-Sna mRNA by in situ hybridization. Picture 2 : Same as picture 1 but in a posterior view. In this orientation, the activation of Ci-Sna expression in the nuclei of the B7.5 blastomeres is clearly visualized.

Conversion to Aniseed format of Ghost database entry from Fujiwara S et al., Mech Dev. 2002; 112Gene name: 115-127. Original annotation: . Expression pattern: Muscle Mesenchyme Nervecord

Expression of Snail at the early gastrula stage. Graphs showing the percentage of embryos with lateral (red) or ectopic (blue) expression of Snail Asterisks mark cells with medial or lateral expression following the same colour code.

Vegetal view of a 110-cell stage wild type probed for Ci-Snail expression.

Vegetal view of a 110-cell stage wild type embryo probed for snail expression.

Original annotation :
Gene name :Snail.
Expression pattern : A8.5, A8.6, A8.13, A8.14, A8.15, A8.16, B8.5, B8.7, B8.8, B7.5, B7.7, B8.15, B8.16.
Comment : The expression in A8.5, A8.6, A8.13 and A8.14 is very weak.

Aniseed annotation : according to the picture, it seems that there is no expression in B7.5 cell pair.

Vegetative view of a 110-cell stage embryo, showing expression of Ci-snail.

Vegetal view of an early gastrula embryo probed for Ci-Snail expression.

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 :Snail.
Expression pattern : A8.5, A8.6, A8.13, A8.14, A8.15, A8.16, B8.5, B8.7, B8.8, B7.5, B7.7, B8.15, B8.16.
Comment : The expression in the notochord lineages A8.5, A8.6, A8.13 and A8.14 is barely detectable.

Vegetal pole view of 110-cell stage embryos probed for Ci-Snail expression in which the b6.5 blastomere was ablated on the right side. 27/43 embryos showed complete loss of Snail in the lateral A-line neural plate, while 13/43 showed a partial loss only. Control sibling embryo shows the wild type pattern of the gene.

To facilitate data mining, the expression pattern is described as lost expression in the A-line when b6.5 derivatives are ablated.

Vegetal view of a WT Ciona robusta 112-cell stage embryo probed for Snail expression.
Expression is observed in A8.6, A8.6, A8.13, A8.14, A8.15, A8.16, B8.5, B8.6, B8.7, B8.8, B8.15, B8.16 and B7.7 blastomere pairs.
In schematic representation of gene expression, notochord precursors are shown in red. Blastomeres with gene expression are marked by dots.

Vegetal view of a 110 cell stage wild type embryo probed for Ci-sna expression. On this picture an intense expression is observed in muscle/trunk mesenchyme precursors and in the progenitors of the lateral ependymal cells, A8.15 and A8.16.Weaker staining is observed in the primary lineage notochord precursor cells.

Vegetative view of Snail espression at the early gastrula stage. Original annotation : Snail is expressed in the B-line muscle lineages (black bracket), and in A8.15 and A8.16 neural lineages (lilac, yellow, blue and orange arrowheads respectively correspond to A8.7, A8.8, A8.15 and A8.16 neural lineages)

Original annotation: ISH for Neurog (red) and Snail (green). Scale bar, 25 μm. Box enlarged in d′ showing low levels of Snail expression in BTN progenitor.
Aniseed annotation: Expression of snail in neural plate border cells.

Vegetal view of a WT Ciona robusta mid gastrula embryo probed for snail expression (green) and electroporated with Msx>LacZ (red). The dashed area is enlarged in a′.
Author comment : Snail expression is revealed in the bipolar tail neuron progenitors (b9.36 cells, arrowheads).
Scale bar, 25 μm

Animal view of a mid gastrula embryo for 02928 expression. 02928 encodes a zinc finger transcription factor.

Original annotation : expression was detected in the precursor cells of the muscle (mu) and a subset of the nerve cord cells (nc).

Aniseed annotation : precursors of nerve cord are A9.32, A9.29, A9.15, A9.13, b9.37 cell pairs. According to the picture, it seems that the staining is localized in A9.32 and b9.37 cell pairs

Vegetal view of a gastrula stage wild type embryo probed for Ci-Snail expression.

Neural plate view of a mid gastrula stage wild type embryo probed for Ci-sna expression. Staining appears in the muscle precursors that lies between the posterior and anterior patches as well as in the a8.25 blastomere pair and CNS precursors.

Conversion to Aniseed format of Ghost database entry from Fujiwara S et al., Mech Dev. 2002; 112Gene name: 115-127. Original annotation: . Expression pattern: Muscle Nervecord

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 :Snail.
Expression pattern : muscle, unidentifiable.
Comment : Nervous system: A9.29, A9.30, A9.32, a9.49, a9.50, b9.34, b9.37, b9.38

Vegetal view of a late gastrula stage embryo probed for Ci-Snail expression. Ci-snail was expressed in a-line neural cells, b-line neural cells and A-line nerve cord cells, more precisely in A9.29, A9.30, A9.32, a9.49, a9.50, b9.33, b9.37, b9.38.

Vegetal view of a WT Ciona robusta 6-row neural plate stage embryo probed for Snail expression and counterstained with DAPI.
Snail expression is observed in a9.49, A9.29, A9.30, A9.31 and A9.32 blastomers, and in tail muscles.
Original annotation: Snail is expressed downstream of Nodal in the row III/column 3 precursor at the 6-row neural plate stage.
In the drawing, Snail expression is indicated by black dots, with weaker expression represented by grey dots. The a-lineage neural plate cells that generate the CNS are coloured in red.

Vegetal view of a WT Ciona robusta 6-row neural plate stage embryo probed for Snail expression.
Snail expression is observed in a9.49 (column 3 of neural plate row III, pink arrowhead), A9.32, A9.29 and tail muscle.
n=total number of embryos analysed.
Second image is a HD picture of the same 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 :Snail.
Expression pattern : muscle

Dorsal view of an initial tailbud showing GFP expression in electroporated embryo with the ci-sna -3.2kb codon 15 region driving GFP.

Most of the intense fluorescence that is out of the plane focus in the posterior of the embryo corresponds to expression in tail muscles. To the right of the arrow lie cells that have converged at the dorsal midline and which derive from A8.15 and A8.16. Fluorescing cells that lie anterior to the arrow correspond to progenitors of the sensory vesicle derived from a8.25 that have not yet converged at the dorsal midline.

Aniseed annotation: expression in the TVCs and Notochord was added as it is detected in PMID 9501022.

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 :Snail.
Expression pattern : muscle

Lateral and dorsal views of a stained embryo showing Ci-sna expression in tail muscles (arrowhead). There is no detectable staining in the notochord (arrow)

Sagittal view of a mid tail bud stage embryo electroporated with a construct containing the truncated B4.1 enhancer Ci-Sna -482bp to -168bp attached to the Ci-Bra minimal promoter driving the expression of a lacZ reporter gene. Expression is obtained in the trunk mesenchyme, but staining is nearly abolished in the primary muscles.

Lateral view of an early tailbud stage showing expression of Ci-sna fusion genes in electroporated embryo with a construct which include the 3,2kb of the 5 flanking sequence of the Ci-Snail promoter region, as well as 1,3kb of genomic DNA sequences , including a short 5 UTR, exon 1 (58bp), a 1,1kb intron, and the first 50-bp of exon 2. This picture shows expression in the two rows of A8.15 and A8.16 derived ependymal cells in the spinal cord. The row indicated by white arrow appear fainter since it is out of the plane of focus.

Aniseed annotation: activity of this construct in the TVCs and Notochord cannot be seen on this picture, but is indicated in Fig. 2 of PMID 9501022.

Sagittal view of a mid-tail bud stage embryo, electroporated with a lacZ transgene containing a mutated version of ci-sna -618 -356 (262bp Ci-Sna) with point mutations in the T2 and T3 T-Box motifs, abalting the function of these sites. Only very faint residual staining is detected in the tail muscles (short arrow) and mesenchyme (long arrow).

Sagittal view of a mid-tail bud stage embryo, electroporated with a lacZ transgene containing a mutated Ci-Sna -618 -360 (262bp Ci-Sna) enhancer with point mutations in the T2 and T3 T-Box motifs. The G residues of position 5 within the two T-Box motifs were mutagenized G --> A. There is a complete loss of staining.

Sagittal view of a mid-tail bud stage embryo, electroporated with a lacZ transgene containing the 3,2kb of the 5' flanking Ci-Sna promoter sequence, with a small internal deletion within the full length promoter wich removes the 262bp-enhancer. This results in reduced staining in tail-bud embryos. Note the loss of the sensory vesicle staining, which may indicate an overall reduction of enhancer activity rather than a specific loss of muscle and mesenchyme enhancer activity.

No picture available. Mid tailbud stage embryo electroporated with a construct which include the sequences from -3,2kb to -68bp of the Ci-Sna 5' falnking sequences attached to the heterologous Ci-Bra basal promoter. In this construct the Ci-Sna 5 DNA fragment is placed in the opposite (anti) orientation as the direction of Ci-Bra/LacZ transcription.

This construct is transcriptionaly inactive.

Sagittal view of a mid-tail bud stage embryo, electroporated with a lacZ transgene containing the 3,2kb of the 5' flanking Ci-Sna promoter sequence. Intense staining is observerd in the tail muscles, the trunk mesenchyme, the adult muscle precursor cells and the Central Nervous System. Staining is additionally detected in the TVCs and notochord, though it cannot be seen on this picture.

Picture 1 : Sagittal view of a mid-tail bud stage embryo, electroporated with a lacZ transgene containing the -2,2kb of the 5' flanking Ci-Sna promoter sequence. Staining is observerd in the cerebral vesicle, the trunk ventral cells, the primary and secondary muscles.

Picture 2 : Same as picture 1 except that this electroporated embryo exhibits additional aspects of the Ci-Sna expression pattern. Staining is observed in the lateral ependymal cell in the neural tube and notochord.

: the difference between the two pictures reflects the mosaicism obtained in electroporation experiments.

Lateral view of a mid tailbud stage showing GFP expression driven by Ci-sna -3.2kb to codon 15 in electroporated embryo. fluorescence is detected in the muscle cells of the tail, in the trunk mesenchyme cells, in the lateral ependymal cells and in a portion of the anterior sensory vesicle.

Aniseed annotation: expression in the TVCs and Notochord was added as it is detected in PMID 9501022.

No PIcture available. Staining is the same as in the -3.2kb-codon15 region: sensory vesicle, trunk ventral cells, primary and secondary muscles, lateral ependymal cells in the neural tube and notochord.

No picture available. Mid tailbud stage embryo electroporated with a construct which include the sequences from -1,7kb to -738bp of the Ci-Sna 5 regulatory region attached to the heterologous Ci-Bra basal promoter. In this construct the Ci-Sna 5 DNA fragment is placed in the same (syn) orientation as the direction of Ci-Bra/LacZ transcription. This construct is inactive.

No picture available. Mid tailbud stage embryo electroporated with a construct which include the sequences from -1,7kb to -738bp of the Ci-Sna 5 regulatory region attached to the heterologous Ci-Bra basal promoter. In this construct the Ci-Sna 5 DNA fragment is placed in the opposite (anti) orientation as the direction of Ci-Bra/LacZ transcription. This construct is inactive.

Sagittal view of a mid tail bud stage embryo electroporated with a construct containing a Ci-Sna/Ci-fkh fusion promoter. The Ci-sna promoter containing 262bp extend from -618bp to -356bp, while the Ci-fkh promoter regions extends to approx -180bp upstream of the transcribed region. The expression is detected in the tail muscles and trunk mesenchyme.

Aniseed annotation: As the B7.5 tail muscle lineage is labelled, expression is also probably detected in the TVCs.

Sagittal view of a late tail bud, electroporated with a construct containing the "full-length" Ci-Sna/LacZ transgene lacking an internal 1.5kb EcoRI fragment. This transgene directs weak expression in secondary lineage, including lateral ependymal cells in the neural tube (medium arrow) and secondary muscle cells that drive from the A7.8 blastomere (long arrow). Staining is also observed in the cerebral vesicle (short arrow).

NO picture available. Mid tailbud stage embryo electroporated with the construct which include the sequences from -3,2kb to -68bp of the Ci-Sna 5' flanking sequences attached to the Ci-Bra basal promoter driving LacZ. The Ci-Sna 5 DNA fragment is placed in the same (syn) orientation as the direction of Ci-Bra/LacZ transcription.

This construct directs the full Ci-sna expression pattern as well as notochord expression.

No picture available. At the Mid tailbud stage, no activity is detected. In this construct the Ci-Sna 5' DNA fragment is placed in the opposite (anti) orientation as the direction of Ci-Bra/LacZ transcription.

No picture available.In this construct the Ci-Sna 5 DNA fragment is placed in the same (syn) orientation as the direction of Ci-Bra/LacZ transcription. Mid tailbud stage embryo electroporated with this construct mediate expression in the primary muscle lineages, TVCs (not sure) and B-line mesenchyme, but is inactive in the secondary muscle lineages, CNS and notochord.

No picture available. Mid-tailbud stage embryo, electroporated with a lacZ transgene containing the -321bp of the 5' flanking Ci-Sna promoter sequence. No activity.

No PIcture available. No staining detected.

No picture available. Mid-tailbud stage embryo, electroporated with a lacZ transgene containing the -68bp of the 5' flanking Ci-Sna promoter sequence, show no staining.

No picture available. Embryos were electroporated with a construct which include the sequences from -691bp to -168bp of the Ci-Sna 5 regulatory region attached to the heterologous Ci-Bra basal promoter. the Ci-Sna 5 DNA fragment is placed in the same (syn) orientation as the direction of Ci-Bra/LacZ transcription.

This construct mediates expression in the primary B4.1 lineages, in the primary muscles and mesenchyme , but not in the trunk ventral cells.

No picture available. This construct is inactive.

No picture available. Embryos were electroporated with a construct which include the sequences from -691bp to -168bp of the Ci-Sna 5 regulatory region attached to the heterologous Ciona forkhead promoter region. In this construct the Ci-Sna 5 DNA fragment is placed in the opposite (Anti) orientation as the direction of Ci-fkh/LacZ transcription. This construct directs expression in the primary muscles and mesenchyme but not in the TVCs.

Embryos electroporated with this construct which contains 446 bp of ci-sna promoter sequence show no lacZ staining.

No picture available. Embryos electroporated with the sna 531bp construct show weak expression (20% of embryos) in primary muscles and trunk mesenchyme.

lateral view of a midtail bud, electroporated with a construct containing the 737bp Ci-Sna promoter driving the expression of a lacZ reporter gene containing a nuclear localization signal. Staining is observed in the primary muscle and trunk mesenchyme.

No picture availableMid tail bud, electroporated with a construct which include the sequence from -737bp to -1bp, which as s mutation on the 3rd distal AC-core E-Box. Ac-core E-box -> TT-core E-box. This construct, like its parental non mutated construct, directs strong expression in primary muscles and trunk mesenchyme.

No picture available. Embryos were electroporated with a construct which include the sequence from -650bp to +87bp of Sna gene, which has a mutation on the 2nd distal AC-core E-Box (E6 site). Ac-core E-box -> TT-core E-box. This construct directs weak expression in primary muscles and trunk mesenchyme. Only 31% of embryos labelled.

No picture available. Embryos were electroporated with a construct which include the sequence from -650bp to +87bp of Sna gene, which has a mutation on the 1st distal AC-core E-Box (E7 site). Ac-core E-box -> TT-core E-box. This construct directs slightly weaker expression in primary muscles and trunk mesenchyme than the non mutated version. 65% of embryos labelled.

No picture available. Embryos were electroporated with a construct which include the sequence from -650bp to +87bp of Sna gene, which has a mutation on the 2nd and third distal AC-core E-Box (E6, E5 sites). Ac-core E-box -> TT-core E-box. This construct directs weak expression in primary muscles and trunk mesenchyme. Only 22% of embryos labelled.

No picture available. Embryos were electroporated with a construct which include the sequence from -650bp to +87bp of Sna gene, which has a mutation in the 1st and 2nd distal AC-core E-Boxes (E7, E6 site). Ac-core E-box -> TT-core E-box. This construct directs weak expression in primary muscles and trunk mesenchyme. Only 22% of embryos labelled.

Lateral view of a mid tailbud embryo, electroporated with a construct containing the 737bp Ci-Sna promoter which all three AC-core E-boxes have been changed to TT-cores. Intense staining is seen in the neural tube (arrows). Some embryos also exhibits expression in the secondary muscle and the sensory vesicle. While expression in primary muscles is abolished, some embryos continue to show staining in trunk mesenchyme.

Lateral view of a mid tailbud stage showing expression of Ci-sna fusion genes in electroporated embryos with a construct which include the 3,2kb of the 5 flanking sequence of the Ci-Snail promoter region, as well as 1,3kb of genomic DNA sequences , including a short 5 UTR, exon 1 (58bp), a 1,1kb intron, and the first 50-bp of exon 2. This picture shows expression in all the muscles of the tail and in the trunk mesenchyme cells.The white arrows indicates staining in the dorsal portion of the cerebral vesicle in descendance of a8.25. The black arrow indicates four B7.5 derived 'trunk ventral cells' which are the progenitors of adult muscle cells.

Aniseed annotation: expression in the lateral ependymal cells and Notochord is detected with this construct although this cannot be clearly seen on the picture (see PMID 9501022).

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 :Snail.
Expression pattern : No distinct zygotic signal was detected

lateral view of a late tailbud embryo, electroporated with a construct containing the -465bp to +1.3 kb Ci-Sna region driving the expression of a lacZ reporter gene containing a nuclear localization signal. This is the smallest transgene that gives the full Ci-Sna expression pattern. Staining is observed in primary muscle , TVCs, mesenchyme sensory vesicle, notochord and in A-line secondary muscles and lateral ependymal cells in the neural tube. Due to mosaicism, only part of the full expression of the transgene is seen on the picture.

Sagittal view of a mid tailbud stage embryo electroporated with a construct containing the 540bp Ci-Sna promoter placed in syn to the Ci-fkh promoter region driving the expression of a lacZ reporter gene. The particular embryo shown here lacks staining in the trunk mesenchyme, however other embryos exhibits staining in the mesenchyme. The anterior primary muscles consistently exhibits staining.