1021C Poster - 16. Techniques and technology
Saturday April 09, 1:30 PM - 3:30 PM

Authors:
Christopher Sottolano ¹; Nicole Revaitis ¹; Anthony Geneva ^1,2; Nir Yakoby ^1,2

Affiliations:
1) Center for Computational and Integrative Biology, Rutgers University, Camden, NJ; 2) Department of Biology, Rutgers University, Camden, NJ

Keywords:
g. CRISPR/Cas9; k. next-generation sequencing

The plethora of morphological diversity among Drosophila species presents an opportunity to study the mechanisms underlying the molecular evolution of tissue patterning and morphologies. One of the challenges in investigating these species is that unlike the tremendous number of molecular and genetics tools available for Drosophila melanogaster research, many other species do not have sequenced genomes. In addition, the available molecular tools to manipulate genes are very limited. Here, we focus on D. nebulosa due to the ease of rearing, its differences in tissue patterning and eggshell morphologies, and outstanding organismal behavior when compared to D. melanogaster. As a first step to develop the fly as a new genomic and genetic resource, we generated a high-quality annotated genome assembly of D. nebulosa, using PacBio long-read sequencing. We utilized the assembly to successfully disrupt the white gene via CRISPR/Cas9, but were unable to integrate the Cas9 gene using homology-directed repair. Interestingly, unlike many other Drosophila species, D. nebulosa males null for the white gene did not appear to court females. We conclude, gene disruption via CRISPR/Cas9 genome engineering is a useful tool in D. nebulosa. However, a simple selectable marker which does not impair vision is needed to replace the commonly used white-eye phenotype.