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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Journal ArticleDOI
Design and Generation of Donor Constructs for Genome Engineering in Drosophila.
TL;DR: A protocol for the generation of donor constructs using Golden Gate assembly is described and some key considerations for donor construct design for use in Drosophila are discussed.
Posted ContentDOI
A versatile two-step CRISPR- and RMCE-based strategy for efficient genome engineering in Drosophila
TL;DR: An efficient, two-step strategy to flexibly engineer the fly genome by combining CRISPR with recombinase-mediated cassette exchange (RMCE) is developed, suggesting that any fly lab can engineer their favourite gene for a broad range of applications within about three months.
Journal ArticleDOI
Conditional Synaptic Vesicle Markers for Drosophila.
TL;DR: The development and characterization of four conditional tagged SV markers for Drosophila melanogaster are presented, including evaluation of conditionality, specificity for SV localization, and sensitivity of detection in diverse neuron subtypes.
Journal ArticleDOI
CRISPR Start-Loss: A Novel and Practical Alternative for Gene Silencing through Base-Editing-Induced Start Codon Mutations.
Siyu Chen,Wanhua Xie,Zhiquan Liu,Huanhuan Shan,Mao Chen,Yuning Song,Hao Yu,Liangxue Lai,Zhanjun Li +8 more
TL;DR: A novel and practical alternative method referred to as CRISPR Start-Loss (CRISPR-SL), which eliminates gene expression by utilizing both adenine base editors and cytidine base editors to disrupt the initiation codon (ATG).
Patent
Transgenic non-human vertebrate for the expression of class - switched, fully human, antibodies
TL;DR: In this paper, the authors proposed a method to obtain antigen-specific antibodies that are fully human (i.e., comprising human variable and constant regions) and have undergone recombination, junctional diversification, affinity maturation and isotype switching in vivo in a non-human vertebrate system.
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.