Oocytes of a single individual of Halocynthia roretzi (Aomori, Japan) were fertilized by sperms of an another single individual and development was followed microscopically. Embryos were snapfrozen in liquid nitrogen at the appropriate stage and kept at -80°C. Total RNA was prepared with the AGPC method and the quality was checked with the Bioanalyzer, most 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).

Embryo probed for Hr-Lhx3a expression (genbank accession number for the clone : D38572)

Original Annotation :
Faint signals of transcriptional isoform a were detected in unfertilized or fertilized eggs, with the signals being seen throughout the cell.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : D38572

Embryo probed for Hr-Lhx3b expression (genbank accession number for the clone : AB5007042)

Original Annotation :
No expression of Hr-Lhx3b was detected.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : AB500704

2-cell stage wild type embryo probed for Hr-Lhx3a expression (Genbank accession number for the clone : D38572)

Original Annotation :
Faint WISH signals were also detected in a half of each blastomere in 2-cell stage embryos.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : D38572

2-cell stage wild type embryo probed for Hr-Lhx3b expression (Genbank accession number for the clone : AB500704)

Original Annotation :
No expression detected for Hr-Lhx3b.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : AB500704

4-cell stage wild type embryo probed for Hr-Lhx3a expression (Genbank accession number for the clone : D38572)

Original Annotation :
Faint WISH signals were also detected in a half of each blastomere in 4-cell stage embryos.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : D38572

4-cell stage wild type embryo probed for Hr-Lhx3b expression (Genbank accession number for the clone : AB500704)

Original Annotation :
No expression detected for Hr-Lhx3b.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : AB500704

E8-cell stage wild type embryo probed for Hr-Lhx3a expression (Genbank accession number for the clone : D38572)

Original Annotation :
Faint WISH signals were detected in the animal half of 8-cell stage embryos.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : D38572

8-cell stage wild type embryo probed for Hr-Lhx3b expression (Genbank accession number for the clone : AB500704)

Original Annotation :
No expression detected for Hr-Lhx3b.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : AB500704

Animal, lateral, and vegetal views of a 16-cell stage wild type embryo probed for Hr-Lhx3a expression (Genbank accession number for the clone : D38572).

Original Annotation :
Faint WISH signals were detected in the animal half of 16-cell stage embryos.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : D38572

Animal, lateral, and vegetal views of a 16-cell stage wild type embryo probed for Hr-Lhx3b expression (Genbank accession number for the clone : AB500704).

Original Annotation :
No expression detected for Hr-Lhx3b.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : AB500704

Vegetal view of a 32-cell stage wild type embryo probed for Hr-Lhx3a expression (Genbank accession number for the clone : D38572).

Original Annotation :
No expression was detected.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : D38572

Vegetal view of a 32-cell stage wild type embryo probed for Hr-Lhx3b expression (Genbank accession number for the clone : AB500704).

Original Annotation :
Expression of Lhx3b were detected by WISH from the 32-cell stage onward in the endoderm producing cells, A6.1, A6.3 and B6.1 pairs.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : AB500704

Vegetal view of a 64-cell stage wild type embryo probed for Hr-Lhx3b expression (Genbank accession number for the clone : AB500704).

Original Annotation :
The expression of Lhx3b continues in the endoderm lineage blastomeres (A7.1, A7.2, A7.5, B7.1, B7.2, B7.5) as well as the notochord and mesenchyme blastomeres (A7.3, A7.7, B7.3) at the 64-cell stage.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : AB500704

Vegetal view of a 64-cell stage wild type embryo probed for Hr-Lhx3a expression (Genbank accession number for the clone : D38572).

Original Annotation :
Hr-Lhx3a was not expressed at this stage.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : D38572

Vegetal view of a 110-cell stage wild type embryo probed for Hr-Lhx3a expression (Genbank accession number for the clone : D38572).

Original Annotation :
Hr-Lhx3a was not expressed at this stage.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : D38572

Vegetal view of a 110-cell stage wild type embryo probed for Hr-Lhx3b expression (Genbank accession number for the clone : AB500704).

Original Annotation :
The expression of Lhx3b continues in the endoderm lineage blastomeres (A7.1, A7.2, A7.5, B7.1, B7.2, B7.5) as well as the notochord and mesenchyme blastomeres (A7.3, A7.7, B7.3) at the 110-cell stage.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : AB500704

Vegetal view of a gastrula stage wild type embryo probed for Hr-Lhx3a expression (Genbank accession number for the clone : D38572).

Original Annotation :
Hr-Lhx3a was not expressed at this stage.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : D38572

Vegetal view of a gastrula stage wild type embryo probed for Hr-Lhx3b expression (Genbank accession number for the clone : AB500704).

Original Annotation :
Expression of Lhx3b were detected by WISH from the 32-cell stage onward in the endoderm producing cells, A6.1, A6.3 and B6.1 pairs, and the expression of Lhx3b continues in the endoderm lineage blastomeres (A7.1, A7.2, A7.5, B7.1, B7.2, B7.5) as well as the notochord and mesenchyme blastomeres (A7.3, A7.7, B7.3) at the 64-cell stage. Thereafter, the expression became weaker and disappeared by the neurula stage.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : AB500704

Dorsal view of a neurula stage wild type embryo probed for Hr-Lhx3b expression (Genbank accession number for the clone : AB500704).

Original Annotation :
The expression disappeared by the neurula stage.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : AB500704

Dorsal view of a neurula stage wild type embryo probed for Hr-Lhx3a expression (Genbank accession number for the clone : D38572).
Insets are specimens detected for the transcripts of Hr-Lhx3a by WISH using the probe covering the entire coding region of Hr-Lhx3a.

Original Annotation :
Zygotic expression of Lhx3a becomes detectable by WISH from the neurula stage onward in the cells at the neural fold.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : D38572

Lateral view of an early tailbud stage wild type embryo probed for Hr-Lhx3a expression (Genbank accession number for the clone : D38572).
Insets are specimens detected for the transcripts of Hr-Lhx3a by WISH using the probe covering the entire coding region of Hr-Lhx3a.

Original Annotation :
Zygotic expression of Lhx3a becomes detectable in a part of brain and the visceral ganglion cells at the tailbud stages.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : D38572

Lateral view of an early tailbud stage wild type embryo probed for Hr-Lhx3b expression (Genbank accession number for the clone : AB500704).

Original Annotation :
The expression became weaker and disappeared by the neurula stage.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : AB500704

Lateral view of a mid tailbud I stage wild type embryo probed for Hr-Lhx3b expression (Genbank accession number for the clone : AB500704).

Original Annotation :
The expression became weaker and disappeared by the neurula stage.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : AB500704

Lateral view of a mid tailbud I stage wild type embryo probed for Hr-Lhx3a expression (Genbank accession number for the clone : D38572).

Original Annotation :
Zygotic expression of Lhx3a becomes detectable in a part of brain and the visceral ganglion cells at the tailbud stages. As for the expression of Hr-Lhx3a in the visceral ganglion located between the trunk and tail, detailed analysis was performed. The analysis revealed that at the mid tailbud I stage Hr-Lhx3a seemed to be expressed in the two rows of three cells, these cells are motor neurons.

Aniseed Annotation :
Be careful to the clone linked to the annotation in this experiment: the clones used for the two isoforms Lhxa and Lhxb are not yet in the database. So the same clone was used for annotation of Hr-Lhxa and Hr-Lhxb.
Genbank accession number of the sequence : D38572

Expression of Hr-Lhx3a in the visceral ganglion region of a mid tailbud I stage embryo as revealed by fluorescence WISH. Fluorescence images of the dorsal views of higher magnification of the visceral ganglion region are shown. Anterior is towards the top. Note that the signals were concentrated at the six nuclei.