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...

Descripción completa

Detalles Bibliográficos
Autores principales: 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
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