title: Clustered regulatory interspaced short palindromic repeats (CRISPR)-mediated mutagenesis and phenotype rescue by piggyBac transgenesis in a nonmodel Drosophila species.
abstract: How behavioural diversity emerged in evolution is an unexplored subject in biology. To tackle this problem, genes and circuits for a behaviour need to be determined in different species for phylogenetic comparisons. The recently developed clustered regulatory interspaced short palindromic repeats/CRISPR associated protein9 (CRISPR/Cas9) system made such a challenge possible by providing the means to induce mutations in a gene of interest in any organism. Aiming at elucidating diversification in genetic and neural networks for courtship behaviour, we attempted to generate a genetic tool kit in Drosophila subobscura, a nonmodel species distantly related to the genetic model Drosophila melanogaster. Here we report the generation of yellow (y) and white mutations with the aid of the CRISPR/Cas9 system, and the rescue of the y mutant phenotype by germline transformation of the newly established y mutant fly line with a y(+) -marked piggyBac vector. This successful mutagenesis and transformation in D. subobscura open up an avenue for comprehensive genetic analyses of higher functions in this and other nonmodel Drosophila species, representing a key step toward systematic comparisons of genes and circuitries underlying behaviour amongst species.
authors.affiliation: Division of Neurogenetics, Graduate School of Life Sciences, Tohoku University, Sendai, Japan.|Division for International Advanced Research and Education, Tohoku University, Sendai, Japan.
authors.forename: R
authors.initials: R
authors.lastname: Tanaka
authors.affiliation: Division of Neurogenetics, Graduate School of Life Sciences, Tohoku University, Sendai, Japan.
authors.forename: H
authors.initials: H
authors.lastname: Murakami
authors.affiliation: Division of Neurogenetics, Graduate School of Life Sciences, Tohoku University, Sendai, Japan.
authors.forename: M
authors.initials: M
authors.lastname: Ote
authors.affiliation: Division of Neurogenetics, Graduate School of Life Sciences, Tohoku University, Sendai, Japan.
authors.forename: D
authors.initials: D
authors.lastname: Yamamoto
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D001483:Base Sequence; D064112:Clustered Regularly Interspaced Short Palindromic Repeats; D004330:Drosophila / Q000145:classification / Q000235:genetics* / Q000378:metabolism; D029721:Drosophila Proteins / Q000235:genetics* / Q000378:metabolism; D005136:Eye Proteins / Q000235:genetics* / Q000378:metabolism; D005260:Female; D018014:Gene Transfer Techniques; D008297:Male; D016296:Mutagenesis; D010641:Phenotype
chemical_list: D018528:ATP-Binding Cassette Transporters; D029721:Drosophila Proteins; D005136:Eye Proteins; C059254:w protein, Drosophila; C050680:y protein, Drosophila
keywords: genome editing; germline transformation; pigmentation mutations; visible marker