Photoreception and vision-related gene characterisation has been studied in sea urchinsin the past. However, not in Paracentrotus lividus, a fast developing and local species. The aims of this project include: the characterisation of PlOpsin4 in late larval and post-metamorphic stages, using CRISPR/Cas9 technology; immuno-detection of PlOpsin4; and the development of a protocol to culture P. lividusfrom fertilisation through to metamorphosis. The results were unfavourable for the objective involving a knockout of PlOpsin4 with CRISPR/Cas9, as no mutation was induced in the gene of interest. Larval cultures successfully managed to develop through all larval stages and metamorphosis, resulting in juvenile sea urchins. De novo characterisation of PlOpsin4 through immunolabeling was achieved effectively with signal detected at the basal position of primary tube feet in 8 arm larvae rudiment and juveniles and on the rim of tube feet discs in juveniles and adults. The issues encountered with CRISPR/Cas9 technology are thought usual when developing a new protocol and will be revised. Immunohistochemistry as a method to validate PlOpsin4 knockouts will be refined. Larval culturing will be improved to attain better survival rates throughout all stages. Overall, the partial characterisation of PlOpsin4 and a better understanding of photoreception was accomplished.
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