Cargando…
Highly efficient DNA-free gene disruption in the agricultural pest Ceratitis capitata by CRISPR-Cas9 ribonucleoprotein complexes
The Mediterranean fruitfly Ceratitis capitata (medfly) is an invasive agricultural pest of high economic impact and has become an emerging model for developing new genetic control strategies as an alternative to insecticides. Here, we report the successful adaptation of CRISPR-Cas9-based gene disrup...
Autores principales: | , , , , , , , , , , , , , , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Nature Publishing Group UK
2017
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5577161/ https://www.ncbi.nlm.nih.gov/pubmed/28855635 http://dx.doi.org/10.1038/s41598-017-10347-5 |
_version_ | 1783260298467082240 |
---|---|
author | Meccariello, Angela Monti, Simona Maria Romanelli, Alessandra Colonna, Rita Primo, Pasquale Inghilterra, Maria Grazia Del Corsano, Giuseppe Ramaglia, Antonio Iazzetti, Giovanni Chiarore, Antonia Patti, Francesco Heinze, Svenia D. Salvemini, Marco Lindsay, Helen Chiavacci, Elena Burger, Alexa Robinson, Mark D. Mosimann, Christian Bopp, Daniel Saccone, Giuseppe |
author_facet | Meccariello, Angela Monti, Simona Maria Romanelli, Alessandra Colonna, Rita Primo, Pasquale Inghilterra, Maria Grazia Del Corsano, Giuseppe Ramaglia, Antonio Iazzetti, Giovanni Chiarore, Antonia Patti, Francesco Heinze, Svenia D. Salvemini, Marco Lindsay, Helen Chiavacci, Elena Burger, Alexa Robinson, Mark D. Mosimann, Christian Bopp, Daniel Saccone, Giuseppe |
author_sort | Meccariello, Angela |
collection | PubMed |
description | The Mediterranean fruitfly Ceratitis capitata (medfly) is an invasive agricultural pest of high economic impact and has become an emerging model for developing new genetic control strategies as an alternative to insecticides. Here, we report the successful adaptation of CRISPR-Cas9-based gene disruption in the medfly by injecting in vitro pre-assembled, solubilized Cas9 ribonucleoprotein complexes (RNPs) loaded with gene-specific single guide RNAs (sgRNA) into early embryos. When targeting the eye pigmentation gene white eye (we), a high rate of somatic mosaicism in surviving G0 adults was observed. Germline transmission rate of mutated we alleles by G0 animals was on average above 52%, with individual cases achieving nearly 100%. We further recovered large deletions in the we gene when two sites were simultaneously targeted by two sgRNAs. CRISPR-Cas9 targeting of the Ceratitis ortholog of the Drosophila segmentation paired gene (Ccprd) caused segmental malformations in late embryos and in hatched larvae. Mutant phenotypes correlate with repair by non-homologous end-joining (NHEJ) lesions in the two targeted genes. This simple and highly effective Cas9 RNP-based gene editing to introduce mutations in C. capitata will significantly advance the design and development of new effective strategies for pest control management. |
format | Online Article Text |
id | pubmed-5577161 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2017 |
publisher | Nature Publishing Group UK |
record_format | MEDLINE/PubMed |
spelling | pubmed-55771612017-09-01 Highly efficient DNA-free gene disruption in the agricultural pest Ceratitis capitata by CRISPR-Cas9 ribonucleoprotein complexes Meccariello, Angela Monti, Simona Maria Romanelli, Alessandra Colonna, Rita Primo, Pasquale Inghilterra, Maria Grazia Del Corsano, Giuseppe Ramaglia, Antonio Iazzetti, Giovanni Chiarore, Antonia Patti, Francesco Heinze, Svenia D. Salvemini, Marco Lindsay, Helen Chiavacci, Elena Burger, Alexa Robinson, Mark D. Mosimann, Christian Bopp, Daniel Saccone, Giuseppe Sci Rep Article The Mediterranean fruitfly Ceratitis capitata (medfly) is an invasive agricultural pest of high economic impact and has become an emerging model for developing new genetic control strategies as an alternative to insecticides. Here, we report the successful adaptation of CRISPR-Cas9-based gene disruption in the medfly by injecting in vitro pre-assembled, solubilized Cas9 ribonucleoprotein complexes (RNPs) loaded with gene-specific single guide RNAs (sgRNA) into early embryos. When targeting the eye pigmentation gene white eye (we), a high rate of somatic mosaicism in surviving G0 adults was observed. Germline transmission rate of mutated we alleles by G0 animals was on average above 52%, with individual cases achieving nearly 100%. We further recovered large deletions in the we gene when two sites were simultaneously targeted by two sgRNAs. CRISPR-Cas9 targeting of the Ceratitis ortholog of the Drosophila segmentation paired gene (Ccprd) caused segmental malformations in late embryos and in hatched larvae. Mutant phenotypes correlate with repair by non-homologous end-joining (NHEJ) lesions in the two targeted genes. This simple and highly effective Cas9 RNP-based gene editing to introduce mutations in C. capitata will significantly advance the design and development of new effective strategies for pest control management. Nature Publishing Group UK 2017-08-30 /pmc/articles/PMC5577161/ /pubmed/28855635 http://dx.doi.org/10.1038/s41598-017-10347-5 Text en © The Author(s) 2017 Open Access 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/. |
spellingShingle | Article Meccariello, Angela Monti, Simona Maria Romanelli, Alessandra Colonna, Rita Primo, Pasquale Inghilterra, Maria Grazia Del Corsano, Giuseppe Ramaglia, Antonio Iazzetti, Giovanni Chiarore, Antonia Patti, Francesco Heinze, Svenia D. Salvemini, Marco Lindsay, Helen Chiavacci, Elena Burger, Alexa Robinson, Mark D. Mosimann, Christian Bopp, Daniel Saccone, Giuseppe Highly efficient DNA-free gene disruption in the agricultural pest Ceratitis capitata by CRISPR-Cas9 ribonucleoprotein complexes |
title | Highly efficient DNA-free gene disruption in the agricultural pest Ceratitis capitata by CRISPR-Cas9 ribonucleoprotein complexes |
title_full | Highly efficient DNA-free gene disruption in the agricultural pest Ceratitis capitata by CRISPR-Cas9 ribonucleoprotein complexes |
title_fullStr | Highly efficient DNA-free gene disruption in the agricultural pest Ceratitis capitata by CRISPR-Cas9 ribonucleoprotein complexes |
title_full_unstemmed | Highly efficient DNA-free gene disruption in the agricultural pest Ceratitis capitata by CRISPR-Cas9 ribonucleoprotein complexes |
title_short | Highly efficient DNA-free gene disruption in the agricultural pest Ceratitis capitata by CRISPR-Cas9 ribonucleoprotein complexes |
title_sort | highly efficient dna-free gene disruption in the agricultural pest ceratitis capitata by crispr-cas9 ribonucleoprotein complexes |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5577161/ https://www.ncbi.nlm.nih.gov/pubmed/28855635 http://dx.doi.org/10.1038/s41598-017-10347-5 |
work_keys_str_mv | AT meccarielloangela highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT montisimonamaria highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT romanellialessandra highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT colonnarita highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT primopasquale highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT inghilterramariagrazia highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT delcorsanogiuseppe highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT ramagliaantonio highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT iazzettigiovanni highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT chiaroreantonia highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT pattifrancesco highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT heinzesveniad highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT salveminimarco highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT lindsayhelen highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT chiavaccielena highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT burgeralexa highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT robinsonmarkd highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT mosimannchristian highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT boppdaniel highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes AT sacconegiuseppe highlyefficientdnafreegenedisruptionintheagriculturalpestceratitiscapitatabycrisprcas9ribonucleoproteincomplexes |