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.

Vegetal view of a WT Ciona robusta neurula embryo probed for Trp expression.
Trp expression is observed in a10.97 and weakly in a 10.98 blastomers.
Original annotation: Trp is expressed in column 3 of neural plate row III daughters, with stronger expression in the posterior cell, a10.97.

Vegetal view of a WT Ciona robusta neurula embryo probed for Trp expression.
Trp expression is observed in a10.98 and a10.97 blastomers.
Original annotation: Trp is expressed in column 3 of neural plate row III daughters, with stronger expression in the posterior cell, a10.97.

Vegetal view of a WT Ciona robusta neurula embryo probed for Trp expression.
Trp expression is observed in a10.98 and a10.97 blastomers from neural plate rows IIIa and IIIb, column 3.

Dorsal view of a WT Ciona robusta mid neurula stage embryo probed for tyrp1/2b expression.
Tyrp1/2b is expressed in a9.49 progeny : a10.97 and a10.98.
Original annotation: Tyrp1/2b hybridization signal was first detected at the middle neurula stage (later to Ci-tyrp1/2a and prior to Ci-tyr).

Dorsal view of a WT Ciona robusta late neurula stage embryo probed for tyrp1/2b expression.
Tyrp1/2b is expressed in a9.49 progeny : a10.97 and a10.98.
Original annotation: Ci-tyr and Ci-tyrp1/2b expression territories were superimposable with that of Ci-tyrp1/2a.

Dorsal view of a Ciona robusta late neurula stage embryo electroporated with tyrp1/2b>mCherry reporter construct. In situ hybridization experiment, using mCherry antisense RNA as probe, was performed in order to reveal the reporter expression at early developmental stages, when fluorescence is still undetectable.
mCherry is expressed in a9.49 progeny : a10.97 and a10.98.
Original annotation: Reporter gene expression perfectly mirrors endogenous genes expression profile.

Early tailbud stage wild type embryo probed for Ci-TYRP1 (citb30d10) expression (no picture available in the article).

Original annotation
Expression was detected in the dorsal part of brain.

Mid tailbud stage wild type embryo probed for Ci-TYRP1 (citb30d10) expression (no picture available in the article).

Original annotation:
Expression was detected in the dorsal part of brain.

Lateral view of a WT Ciona robusta mid tailbud stage embryo probed for tyrp1/2b expression.
Tyrp1/2b is expressed in a9.49 progeny : a10.97 and a10.98 blastomeres that line up along the dorsal midline of the developing neural tube.
Original annotation: Ci-tyr and Ci-tyrp1/2b expression territories were superimposable with that of Ci-tyrp1/2a.

Lateral view of a Ciona robusta mid tailbud stage embryo electroporated with tyrp1/2b>mCherry reporter construct.
mCherry is expressed in a9.49 progeny : a10.97 and a10.98.
Original annotation: Reporter gene expression perfectly mirrors endogenous genes expression profile.

lateral view of embryo.
Original annotation: expression of Tyrp1 in the otolith (green) and ocellus, 10.98 and 10.97 lineage.

Late tailbud stage wild type embryo probed for Ci-TYRP1 (citb30d10) expression (no picture available in the article).

Original annotation
Expression was detected in the dorsal part of brain.

Conversion to aniseed format of DBTGR database entry from Kusakabe team.

Original annotation:
Brain vesicle (pigment cells).

Lateral view of the head of a WT Ciona robusta larva probed for tyrp1/2b expression.
Tyrp1/2b is expressed in otolith an ocellus pigment cells.
Original annotation: Ci-tyr and Ci-tyrp1/2b expression territories were superimposable with that of Ci-tyrp1/2a.

Lateral view of the head of a Ciona robusta larva electroporated with tyrp1/2b>mCherry reporter construct.
mCherry is expressed in otolith an ocellus pigment cells
Original annotation: Reporter gene expression perfectly mirrors endogenous genes expression profile.