Genome engineering of Drosophila with the CRISPR RNA-guided Cas9 nuclease
Scott J. Gratz,Alexander M. Cummings,Jennifer N. Nguyen,Danielle C. Hamm,Laura K. Donohue,Melissa M. Harrison,Jill Wildonger,Kate M. O'Connor-Giles +7 more
TLDR
A bacterial CRISPR RNA/Cas9 system is adapted to precisely engineer the Drosophila genome and it is reported that Cas9-mediated genomic modifications are efficiently transmitted through the germline.Abstract:
We have adapted a bacterial CRISPR RNA/Cas9 system to precisely engineer the Drosophila genome and report that Cas9-mediated genomic modifications are efficiently transmitted through the germline. This RNA-guided Cas9 system can be rapidly programmed to generate targeted alleles for probing gene function in Drosophila.read more
Citations
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Double nicking by RNA-guided CRISPR Cas9 for enhanced genome editing specificity
F. Ann Ran,Patrick D. Hsu,Chie Yu Lin,Jonathan S. Gootenberg,Silvana Konermann,Alexandro E. Trevino,David A. Scott,Azusa Inoue,Shogo Matoba,Yi Zhang,Feng Zhang +10 more
TL;DR: In this paper, an approach that combines a Cas9 nickase mutant with paired guide RNAs to introduce targeted double-strand breaks is described. But the approach is limited to mouse zygotes.
Journal ArticleDOI
High-frequency off-target mutagenesis induced by CRISPR-Cas nucleases in human cells.
Yanfang Fu,Jennifer A Foden,Cyd Khayter,Morgan L. Maeder,Deepak Reyon,J. Keith Joung,Jeffry D. Sander +6 more
TL;DR: It is found that single and double mismatches are tolerated to varying degrees depending on their position along the guide RNA (gRNA)-DNA interface, and off-target cleavage of CRISPR-associated (Cas)9-based RGNs is characterized.
Journal ArticleDOI
CRISPR-Cas systems for editing, regulating and targeting genomes
Jeffry D. Sander,J. Keith Joung +1 more
TL;DR: A modified version of the CRISPR-Cas9 system has been developed to recruit heterologous domains that can regulate endogenous gene expression or label specific genomic loci in living cells, which will undoubtedly transform biological research and spur the development of novel molecular therapeutics for human disease.
Double Nicking by RNA-Guided CRISPR Cas9 for Enhanced Genome Editing Specificity
F. Ann Ran,Patrick D. Hsu,Chie Yu Lin,Jonathan S. Gootenberg,Silvana Konermann,Alexandro E. Trevino,David A. Scott,Azusa Inoue,Shogo Matoba,Yi Zhang,Feng Zhang +10 more
TL;DR: It is demonstrated that using paired nicking can reduce off-target activity by 50- to 1,500-fold in cell lines and to facilitate gene knockout in mouse zygotes without sacrificing on-target cleavage efficiency.
Journal ArticleDOI
Easy quantitative assessment of genome editing by sequence trace decomposition
TL;DR: TIDE, a method that requires only a pair of PCR reactions and two standard capillary sequencing runs to identify the major induced mutations in the projected editing site and accurately determines their frequency in a cell population, is presented.
References
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Journal ArticleDOI
A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity.
Martin Jinek,Krzysztof Chylinski,Krzysztof Chylinski,Ines Fonfara,Michael H. Hauer,Jennifer A. Doudna,Emmanuelle Charpentier +6 more
TL;DR: This study reveals a family of endonucleases that use dual-RNAs for site-specific DNA cleavage and highlights the potential to exploit the system for RNA-programmable genome editing.
Journal ArticleDOI
Multiplex Genome Engineering Using CRISPR/Cas Systems
Le Cong,Le Cong,F. Ann Ran,F. Ann Ran,David M. Cox,David M. Cox,Shuailiang Lin,Shuailiang Lin,Robert P. J. Barretto,Naomi Habib,Patrick D. Hsu,Patrick D. Hsu,Xuebing Wu,Wenyan Jiang,Luciano A. Marraffini,Feng Zhang +15 more
TL;DR: The type II prokaryotic CRISPR (clustered regularly interspaced short palindromic repeats)/Cas adaptive immune system has been shown to facilitate RNA-guided site-specific DNA cleavage as discussed by the authors.
Multiplex Genome Engineering Using CRISPR/Cas Systems
Le Cong,F. A. Ran,David Benjamin Turitz Cox,Shuailiang Lin,Robert P. J. Barretto,Naomi Habib,Patrick D. Hsu,Xuebing Wu,Wenyan Jiang,Luciano A. Marraffini,Feng Zhang +10 more
TL;DR: Two different type II CRISPR/Cas systems are engineered and it is demonstrated that Cas9 nucleases can be directed by short RNAs to induce precise cleavage at endogenous genomic loci in human and mouse cells, demonstrating easy programmability and wide applicability of the RNA-guided nuclease technology.
Journal ArticleDOI
RNA-Guided Human Genome Engineering via Cas9
Prashant Mali,Luhan Yang,Kevin M. Esvelt,John Aach,Marc Güell,James E. DiCarlo,Julie E. Norville,George M. Church,George M. Church +8 more
TL;DR: The type II bacterial CRISPR system is engineer to function with custom guide RNA (gRNA) in human cells to establish an RNA-guided editing tool for facile, robust, and multiplexable human genome engineering.
Journal ArticleDOI
CRISPR provides acquired resistance against viruses in prokaryotes
Rodolphe Barrangou,Christophe Fremaux,Hélène Deveau,Melissa Richards,Patrick Boyaval,Sylvain Moineau,Dennis A. Romero,Philippe Horvath +7 more
TL;DR: It is found that, after viral challenge, bacteria integrated new spacers derived from phage genomic sequences, and CRISPR provided resistance against phages, and resistance specificity is determined by spacer-phage sequence similarity.