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Generation of heritable germline mutations in the jewel wasp Nasonia vitripennis using CRISPR/Cas9

Ming Li, Lauren Yun Cook Au, Deema Douglah, Abigail Chong, Bradley J. White, Patrick M. Ferree, Omar S. Akbari
doi: https://doi.org/10.1101/096578
Ming Li
1Department of Entomology and Riverside Center for Disease Vector Research, Institute for Integrative Genome Biology, University of California, Riverside, Riverside, CA 92521, USA
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Lauren Yun Cook Au
2W.M. Keck Science Department, Claremont McKenna, Pitzer and Scripps Colleges, 925. Mills Avenue, Claremont, CA 91711, USA
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Deema Douglah
2W.M. Keck Science Department, Claremont McKenna, Pitzer and Scripps Colleges, 925. Mills Avenue, Claremont, CA 91711, USA
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Abigail Chong
1Department of Entomology and Riverside Center for Disease Vector Research, Institute for Integrative Genome Biology, University of California, Riverside, Riverside, CA 92521, USA
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Bradley J. White
1Department of Entomology and Riverside Center for Disease Vector Research, Institute for Integrative Genome Biology, University of California, Riverside, Riverside, CA 92521, USA
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Patrick M. Ferree
2W.M. Keck Science Department, Claremont McKenna, Pitzer and Scripps Colleges, 925. Mills Avenue, Claremont, CA 91711, USA
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  • For correspondence: pferree@kecksci.claremont.edu omar.akbari@ucr.edu
Omar S. Akbari
1Department of Entomology and Riverside Center for Disease Vector Research, Institute for Integrative Genome Biology, University of California, Riverside, Riverside, CA 92521, USA
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  • For correspondence: pferree@kecksci.claremont.edu omar.akbari@ucr.edu
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Abstract

The revolutionary RNA-guided endonuclease CRISPR/Cas9 system has proven to be a powerful tool for gene editing in a plethora of organisms. Here, utilizing this system we developed an efficient protocol for the generation of heritable germline mutations in the parasitoid jewel wasp, Nasonia vitripennis, a rising insect model organism for the study of evolution, development of axis pattern formation, venom production, haplo-diploid sex determination, and host–symbiont interactions. To establish CRISPR-directed gene editing in N. vitripennis, we targeted a conserved eye pigmentation gene cinnabar, generating several independent heritable germline mutations in this gene. Briefly, to generate these mutants, we developed a protocol to efficiently collect N. vitripennis eggs from a parasitized flesh fly pupa, Sarcophaga bullata, inject these eggs with Cas9/guide RNA mixtures, and transfer injected eggs back into the host to continue development. We also describe a flow for screening mutants and establishing stable mutant strains through genetic crosses. Overall, our results demonstrate that the CRISPR/Cas9 system is a powerful tool for genome manipulation in N. vitripennis, with strong potential for expansion to target critical genes, thus allowing for the investigation of a number of important biological phenomena in this organism.

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The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-ND 4.0 International license.
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Posted December 23, 2016.
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Generation of heritable germline mutations in the jewel wasp Nasonia vitripennis using CRISPR/Cas9
Ming Li, Lauren Yun Cook Au, Deema Douglah, Abigail Chong, Bradley J. White, Patrick M. Ferree, Omar S. Akbari
bioRxiv 096578; doi: https://doi.org/10.1101/096578
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Generation of heritable germline mutations in the jewel wasp Nasonia vitripennis using CRISPR/Cas9
Ming Li, Lauren Yun Cook Au, Deema Douglah, Abigail Chong, Bradley J. White, Patrick M. Ferree, Omar S. Akbari
bioRxiv 096578; doi: https://doi.org/10.1101/096578

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