Meeting Abstract
Cephalopods exhibit complex behaviors and have the largest nervous systems among the invertebrates. Their large brains and sophisticated neural networks allow them to interact with and respond to their environment with astounding speed and specificity. Cephalopods, most notably the longfin inshore squid, Doryteuthis (Loligo) pealeii, have and continue to play a critical role in our understanding of neural function. Although the mechanisms that direct their development are only now being approached at the molecular level, as genome and transcriptome resources for cephalopods become available, the time is ripe for developing genome editing technologies, such as CRISPR-Cas9, for use in cephalopods. We chose the gene for the enzyme tryptophan dioxygenase (TDO) to target in our experiments. TDO is an enzyme that is necessary for the production of the ommachrome pigments which color the chromatophores and the retina. Successful editing of TDO with CRISPR Cas9 should result in mosaic embryos with regions lacking pigmentation. In this study, in vitro fertilized D. pealeii embryos were injected with Cas9 preincubated with a CRISPR gRNA targeting TDO and cultured to hatching. Injected embryos developed normally and possessed discrete regions of ¼ to ½ albinism. Partial albino embryos were all positive for INDELs in TDO, suggesting that our CRISPR-Cas9 mutagenesis was successful. This study demonstrates for the first time that genome editing studies and the generation of transgenic animal lines are possible for cephalopods. This work was supported by an MBL Fellowship to KC and an NSF EDGE grant (1827509) to our team.
