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Assessment of distant-site rescue elements for CRISPR toxin-antidote gene drives

Gene drive is a genetic engineering technology that can enable super-mendelian inheritance of specific alleles, allowing them to spread through a population. New gene drive types have increased flexibility, offering options for confined modification or suppression of target populations. Among the mo...

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Autores principales: Chen, Jingheng, Xu, Xuejiao, Champer, Jackson
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9968759/
https://www.ncbi.nlm.nih.gov/pubmed/36860883
http://dx.doi.org/10.3389/fbioe.2023.1138702
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author Chen, Jingheng
Xu, Xuejiao
Champer, Jackson
author_facet Chen, Jingheng
Xu, Xuejiao
Champer, Jackson
author_sort Chen, Jingheng
collection PubMed
description Gene drive is a genetic engineering technology that can enable super-mendelian inheritance of specific alleles, allowing them to spread through a population. New gene drive types have increased flexibility, offering options for confined modification or suppression of target populations. Among the most promising are CRISPR toxin-antidote gene drives, which disrupt essential wild-type genes by targeting them with Cas9/gRNA. This results in their removal, increasing the frequency of the drive. All these drives rely on having an effective rescue element, which consists of a recoded version of the target gene. This rescue element can be at the same site as the target gene, maximizing the chance of efficient rescue, or at a distant site, which allows useful options such as easily disrupting another essential gene or increasing confinement. Previously, we developed a homing rescue drive targeting a haplolethal gene and a toxin-antidote drive targeting a haplosufficient gene. These successful drives had functional rescue elements but suboptimal drive efficiency. Here, we attempted to construct toxin-antidote drives targeting these genes with a distant-site configuration from three loci in Drosophila melanogaster. We found that additional gRNAs increased cut rates to nearly 100%. However, all distant-site rescue elements failed for both target genes. Furthermore, one rescue element with a minimally recoded sequence was used as a template for homology-directed repair for the target gene on a different chromosomal arm, resulting in the formation of functional resistance alleles. Together, these results can inform the design of future CRISPR-based toxin-antidote gene drives.
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spelling pubmed-99687592023-02-28 Assessment of distant-site rescue elements for CRISPR toxin-antidote gene drives Chen, Jingheng Xu, Xuejiao Champer, Jackson Front Bioeng Biotechnol Bioengineering and Biotechnology Gene drive is a genetic engineering technology that can enable super-mendelian inheritance of specific alleles, allowing them to spread through a population. New gene drive types have increased flexibility, offering options for confined modification or suppression of target populations. Among the most promising are CRISPR toxin-antidote gene drives, which disrupt essential wild-type genes by targeting them with Cas9/gRNA. This results in their removal, increasing the frequency of the drive. All these drives rely on having an effective rescue element, which consists of a recoded version of the target gene. This rescue element can be at the same site as the target gene, maximizing the chance of efficient rescue, or at a distant site, which allows useful options such as easily disrupting another essential gene or increasing confinement. Previously, we developed a homing rescue drive targeting a haplolethal gene and a toxin-antidote drive targeting a haplosufficient gene. These successful drives had functional rescue elements but suboptimal drive efficiency. Here, we attempted to construct toxin-antidote drives targeting these genes with a distant-site configuration from three loci in Drosophila melanogaster. We found that additional gRNAs increased cut rates to nearly 100%. However, all distant-site rescue elements failed for both target genes. Furthermore, one rescue element with a minimally recoded sequence was used as a template for homology-directed repair for the target gene on a different chromosomal arm, resulting in the formation of functional resistance alleles. Together, these results can inform the design of future CRISPR-based toxin-antidote gene drives. Frontiers Media S.A. 2023-02-13 /pmc/articles/PMC9968759/ /pubmed/36860883 http://dx.doi.org/10.3389/fbioe.2023.1138702 Text en Copyright © 2023 Chen, Xu and Champer. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Bioengineering and Biotechnology
Chen, Jingheng
Xu, Xuejiao
Champer, Jackson
Assessment of distant-site rescue elements for CRISPR toxin-antidote gene drives
title Assessment of distant-site rescue elements for CRISPR toxin-antidote gene drives
title_full Assessment of distant-site rescue elements for CRISPR toxin-antidote gene drives
title_fullStr Assessment of distant-site rescue elements for CRISPR toxin-antidote gene drives
title_full_unstemmed Assessment of distant-site rescue elements for CRISPR toxin-antidote gene drives
title_short Assessment of distant-site rescue elements for CRISPR toxin-antidote gene drives
title_sort assessment of distant-site rescue elements for crispr toxin-antidote gene drives
topic Bioengineering and Biotechnology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9968759/
https://www.ncbi.nlm.nih.gov/pubmed/36860883
http://dx.doi.org/10.3389/fbioe.2023.1138702
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