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Rationally-engineered reproductive barriers using CRISPR & CRISPRa: an evaluation of the synthetic species concept in Drosophila melanogaster

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Title: Rationally-engineered reproductive barriers using CRISPR & CRISPRa: an evaluation of the synthetic species concept in Drosophila melanogaster
Authors: Waters, AJ
Capriotti, P
Gaboriau, DCA
Papathanos, PA
Windbichler, N
Item Type: Journal Article
Abstract: The ability to erect rationally-engineered reproductive barriers in animal or plant species promises to enable a number of biotechnological applications such as the creation of genetic firewalls, the containment of gene drives or novel population replacement and suppression strategies for genetic control. However, to date no experimental data exist that explores this concept in a multicellular organism. Here we examine the requirements for building artificial reproductive barriers in the metazoan model Drosophila melanogaster by combining CRISPR-based genome editing and transcriptional transactivation (CRISPRa) of the same loci. We directed 13 single guide RNAs (sgRNAs) to the promoters of 7 evolutionary conserved genes and used 11 drivers to conduct a misactivation screen. We identify dominant-lethal activators of the eve locus and find that they disrupt development by strongly activating eve outside its native spatio-temporal context. We employ the same set of sgRNAs to isolate, by genome editing, protective INDELs that render these loci resistant to transactivation without interfering with target gene function. When these sets of genetic components are combined we find that complete synthetic lethality, a prerequisite for most applications, is achievable using this approach. However, our results suggest a steep trade-off between the level and scope of dCas9 expression, the degree of genetic isolation achievable and the resulting impact on fly fitness. The genetic engineering strategy we present here allows the creation of single or multiple reproductive barriers and could be applied to other multicellular organisms such as disease vectors or transgenic organisms of economic importance.
Issue Date: 3-Sep-2018
Date of Acceptance: 17-Aug-2018
URI: http://hdl.handle.net/10044/1/62976
DOI: https://dx.doi.org/10.1038/s41598-018-31433-2
ISSN: 2045-2322
Publisher: Nature Publishing Group
Journal / Book Title: Scientific Reports
Volume: 8
Issue: 1
Copyright Statement: © 2018 The Author(s). This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.
Sponsor/Funder: Commission of the European Communities
Funder's Grant Number: 335724
Keywords: Science & Technology
Multidisciplinary Sciences
Science & Technology - Other Topics
GENE DRIVE SYSTEM
TRANSCRIPTIONAL ACTIVATION
IN-VIVO
GENOME
EXPRESSION
ENHANCERS
EMBRYOS
PHI-C31
DESIGN
SGRNAS
Publication Status: Published
Open Access location: https://www.nature.com/articles/s41598-018-31433-2
Article Number: 13125
Online Publication Date: 2018-09-03
Appears in Collections:National Heart and Lung Institute
Faculty of Medicine
Faculty of Natural Sciences



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