Ciona robusta adults were obtained from the National Bio-Resource Project for Ciona (NBRP, Japan). 50 unperturbed and Foxd-morphant embryos were collected at the 32-, 64-, and 112-cell stages. RNA was extracted using a Dynabeads mRNA DIRECT Purification Kit (Thermo Fischer Scientific) and libraries were made with an Ion Total RNA-Seq kit ver 2 (Thermo Fischer Scientific). The libraries were sequenced with an Ion PGM instrument (Thermo Fischer Scientific).

The purpose of this project is to identify genes positively and negatively regulated by signaling molecules that belongs to the TGFbeta superfamily.

Mid tailbud embryos probed for Ci-AP expression following the injection of Ci-Rga-MO.

Original Annotation : The expression of a marker for the endoderm, Ci-AP, was lost in nine (47%) of 19 knock-down embryos, and severely reduced in three (15%) embryos
First picture : Ci-Rga knock-down embryos with a weaker phenotype that develop into embryos covered by an epidermis layer.
Second picture : Ci-Rga knock-down embryos with severe phenotype that develop into cell aggregates.

View of a mid tailbud stage embryo probed with ALP (endoderm) expression. Expression of tissue-specific differenciation markers in morpholino oligonuncleotide-injected embryos at the stage corresponding to the mid-tailbud stage. Embryos injected with MO against cieg007f18.

Original annotation: the expression of endoderm-specific ALP was detected, suggesting that the manipulated embryos did not merely die.

Lateral view of a mid tailbud stage embryo probed with ALP (endoderm) expression. Expression of tissue-specific differenciation markers in morpholino oligonucleotide-injected embryos at the stage corresponding to the mid tailbud stage.

Original annotation Embryos injected with MO against cilv006h21 as classified as " ball" phenotype: they developped into spherical-shaped cell masses, like balls. In these embryos, detection of the tissue specific markers indicated thaht almost the major tissues were lost or reduced.

Lateral view of a mid tailbud stage embryo probed with Ci-ALP (endoderm) expression. Expression of tissue-specific differenciation markers in morpholino oligonucleotide-injected embryos at the stage corresponding to the mid tailbud stage.

Original annotation Embryos injected with MO against cieg028g04 as classified as " irregular" phenotype: embryos formed an irregular cellular mass. The a/p axis and trunk and tail could not be distinguished.

Lateral view of a mid tailbud stage embryo probed with ALP (endoderm) expression. Expression of tissue-specific differenciation markers in morpholino oligonucleotide-injected embryos at the stage corresponding to the mid tailbud stage.

Original annotation: Embryos injected with MO against cieg009p15 as classified as " irregular" phenotype: embryos formed an irregular cellular mass. The a/p axis and trunk and tail could not be distinguished.

Lateral view of a mid tailbud stage embryo probed with ALP (endoderm) expression. Expression of tissue-specific differenciation markers in morpholino oligonucleotide-injected embryos at the stage corresponding to the mid tailbud stage.

Original annotation: Embryos injected with MO against cieg039g16 is classified as " a/p axis" phenotype: embryos were extended along the anterior-posterior (a/p) axis, but did not form normal tadpoles, with head and tail being indistinguishable.

Lateral view of a mid tailbud stage embryo probed with ALP (endoderm) expression. Expression of tissue-specific differenciation markers in morpholino oligonucleotide-injected embryos at the stage corresponding to the mid tailbud stage.

Original annotation: Embryos injected with MO against cicl017h21 is classified as " a/p axis" phenotype: embryos were extended along the anterior-posterior (a/p) axis, but did not form normal tadpoles, with head and tail being indistinguishable.

Lateral view of a mid tailbud stage embryo probed with Ci-ALP (endoderm) expression. Expression of tissue-specific differenciation markers in morpholino oligonucleotide-injected embryos at the stage corresponding to the mid tailbud stage.

Original annotation: Embryos injected with MO against cieg010p13 is classified as " a/p axis" phenotype: embryos were extended along the anterior-posterior (a/p) axis, but did not form normal tadpoles, with head and tail being indistinguishable.

Lateral view of a mid tailbud stage embryo probed with ALP (endodrem) expression. Expression of tissue-specific differenciation markers in morpholino oligonucleotide-injected embryos at the stage corresponding to the mid tailbud stage.

Original annotation: Embryos injected with MO against ciad033i21 is classified as " flat" phenotype: eggs develop into flattened larvae and they lost the expression of markers for endoderm, notochord and mesenchyme.

Phosphatase alkaline activity staining in a Ciona robusta lavae injected with morpholino oligos against Eph-1.
Staining is observed in the head endoderm and at the tip of the tail, as in WT conditions.
Original comment: The late endoderm marker gene expression was not affected by Eph1 morpholino injection.