RT Journal Article SR Electronic T1 Highly efficient DNA-free gene disruption in the agricultural pest Ceratitis capitata by CRISPR-Cas9 RNPs JF bioRxiv FD Cold Spring Harbor Laboratory SP 127506 DO 10.1101/127506 A1 Angela Meccariello A1 Simona Maria Monti A1 Alessandra Romanelli A1 Rita Colonna A1 Pasquale Primo A1 Maria Grazia Inghilterra A1 Giuseppe Del Corsano A1 Antonio Ramaglia A1 Giovanni Iazzetti A1 Antonia Chiarore A1 Francesco Patti A1 Svenia D. Heinze A1 Marco Salvemini A1 Helen Lindsay A1 Elena Chiavacci A1 Alexa Burger A1 Mark D. Robinson A1 Christian Mosimann A1 Daniel Bopp A1 Giuseppe Saccone YR 2017 UL http://biorxiv.org/content/early/2017/04/18/127506.abstract AB The Mediterranean fruitfly Ceratitis capitata (medfly) is an invasive agricultural pest of high economical impact and has become an emerging model for developing new genetic control strategies as 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 sgRNAs into early embryos. When targeting the eye pigmentation gene white eye (we), we observed a high rate of somatic mosaicism in surviving G0 adults. Germline transmission of mutated we alleles by G0 animals was on average above 70%, with individual cases achieving a transmission rate of 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 Ceratitis capitata will significantly advance the design and development of new effective strategies for pest control management.