Cargando…
The Design and In Vivo Evaluation of Engineered I-OnuI-Based Enzymes for HEG Gene Drive
The homing endonuclease gene (HEG) drive system, a promising genetic approach for controlling arthropod populations, utilises engineered nucleases to spread deleterious mutations that inactivate individual genes throughout a target population. Previous work with a naturally occurring LAGLIDADG homin...
Autores principales: | , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2013
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3769382/ https://www.ncbi.nlm.nih.gov/pubmed/24040217 http://dx.doi.org/10.1371/journal.pone.0074254 |
_version_ | 1782283977885745152 |
---|---|
author | Chan, Yuk-Sang Takeuchi, Ryo Jarjour, Jordan Huen, David S. Stoddard, Barry L. Russell, Steven |
author_facet | Chan, Yuk-Sang Takeuchi, Ryo Jarjour, Jordan Huen, David S. Stoddard, Barry L. Russell, Steven |
author_sort | Chan, Yuk-Sang |
collection | PubMed |
description | The homing endonuclease gene (HEG) drive system, a promising genetic approach for controlling arthropod populations, utilises engineered nucleases to spread deleterious mutations that inactivate individual genes throughout a target population. Previous work with a naturally occurring LAGLIDADG homing endonuclease (I-SceI) demonstrated its feasibility in both Drosophila and Anopheles. Here we report on the next stage of this strategy: the redesign of HEGs with customized specificity in order to drive HEG-induced ‘homing’ in vivo via break-induced homologous recombination. Variants targeting a sequence within the Anopheles AGAP004734 gene were created from the recently characterized I-OnuI endonuclease, and tested for cleavage activity and frequency of homing using a model Drosophila HEG drive system. We observed cleavage and homing at an integrated reporter for all endonuclease variants tested, demonstrating for the first time that engineered HEGs can cleave their target site in insect germline cells, promoting targeted mutagenesis and homing. However, in comparison to our previously reported work with I-SceI, the engineered I-OnuI variants mediated homing with a reduced frequency, suggesting that site-specific cleavage activity is insufficient by itself to ensure efficient homing. Taken together, our experiments take a further step towards the development of a viable HEG-based population control strategy for insects. |
format | Online Article Text |
id | pubmed-3769382 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2013 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-37693822013-09-13 The Design and In Vivo Evaluation of Engineered I-OnuI-Based Enzymes for HEG Gene Drive Chan, Yuk-Sang Takeuchi, Ryo Jarjour, Jordan Huen, David S. Stoddard, Barry L. Russell, Steven PLoS One Research Article The homing endonuclease gene (HEG) drive system, a promising genetic approach for controlling arthropod populations, utilises engineered nucleases to spread deleterious mutations that inactivate individual genes throughout a target population. Previous work with a naturally occurring LAGLIDADG homing endonuclease (I-SceI) demonstrated its feasibility in both Drosophila and Anopheles. Here we report on the next stage of this strategy: the redesign of HEGs with customized specificity in order to drive HEG-induced ‘homing’ in vivo via break-induced homologous recombination. Variants targeting a sequence within the Anopheles AGAP004734 gene were created from the recently characterized I-OnuI endonuclease, and tested for cleavage activity and frequency of homing using a model Drosophila HEG drive system. We observed cleavage and homing at an integrated reporter for all endonuclease variants tested, demonstrating for the first time that engineered HEGs can cleave their target site in insect germline cells, promoting targeted mutagenesis and homing. However, in comparison to our previously reported work with I-SceI, the engineered I-OnuI variants mediated homing with a reduced frequency, suggesting that site-specific cleavage activity is insufficient by itself to ensure efficient homing. Taken together, our experiments take a further step towards the development of a viable HEG-based population control strategy for insects. Public Library of Science 2013-09-10 /pmc/articles/PMC3769382/ /pubmed/24040217 http://dx.doi.org/10.1371/journal.pone.0074254 Text en © 2013 Chan et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited. |
spellingShingle | Research Article Chan, Yuk-Sang Takeuchi, Ryo Jarjour, Jordan Huen, David S. Stoddard, Barry L. Russell, Steven The Design and In Vivo Evaluation of Engineered I-OnuI-Based Enzymes for HEG Gene Drive |
title | The Design and In Vivo Evaluation of Engineered I-OnuI-Based Enzymes for HEG Gene Drive |
title_full | The Design and In Vivo Evaluation of Engineered I-OnuI-Based Enzymes for HEG Gene Drive |
title_fullStr | The Design and In Vivo Evaluation of Engineered I-OnuI-Based Enzymes for HEG Gene Drive |
title_full_unstemmed | The Design and In Vivo Evaluation of Engineered I-OnuI-Based Enzymes for HEG Gene Drive |
title_short | The Design and In Vivo Evaluation of Engineered I-OnuI-Based Enzymes for HEG Gene Drive |
title_sort | design and in vivo evaluation of engineered i-onui-based enzymes for heg gene drive |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3769382/ https://www.ncbi.nlm.nih.gov/pubmed/24040217 http://dx.doi.org/10.1371/journal.pone.0074254 |
work_keys_str_mv | AT chanyuksang thedesignandinvivoevaluationofengineeredionuibasedenzymesforheggenedrive AT takeuchiryo thedesignandinvivoevaluationofengineeredionuibasedenzymesforheggenedrive AT jarjourjordan thedesignandinvivoevaluationofengineeredionuibasedenzymesforheggenedrive AT huendavids thedesignandinvivoevaluationofengineeredionuibasedenzymesforheggenedrive AT stoddardbarryl thedesignandinvivoevaluationofengineeredionuibasedenzymesforheggenedrive AT russellsteven thedesignandinvivoevaluationofengineeredionuibasedenzymesforheggenedrive AT chanyuksang designandinvivoevaluationofengineeredionuibasedenzymesforheggenedrive AT takeuchiryo designandinvivoevaluationofengineeredionuibasedenzymesforheggenedrive AT jarjourjordan designandinvivoevaluationofengineeredionuibasedenzymesforheggenedrive AT huendavids designandinvivoevaluationofengineeredionuibasedenzymesforheggenedrive AT stoddardbarryl designandinvivoevaluationofengineeredionuibasedenzymesforheggenedrive AT russellsteven designandinvivoevaluationofengineeredionuibasedenzymesforheggenedrive |