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
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TLDR
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.Abstract:
Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) systems provide bacteria and archaea with adaptive immunity against viruses and plasmids by using CRISPR RNAs (crRNAs) to guide the silencing of invading nucleic acids. We show here that in a subset of these systems, the mature crRNA that is base-paired to trans-activating crRNA (tracrRNA) forms a two-RNA structure that directs the CRISPR-associated protein Cas9 to introduce double-stranded (ds) breaks in target DNA. At sites complementary to the crRNA-guide sequence, the Cas9 HNH nuclease domain cleaves the complementary strand, whereas the Cas9 RuvC-like domain cleaves the noncomplementary strand. The dual-tracrRNA:crRNA, when engineered as a single RNA chimera, also directs sequence-specific Cas9 dsDNA cleavage. Our 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.read more
Citations
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Journal ArticleDOI
Gene replacements and insertions in rice by intron targeting using CRISPR-Cas9.
TL;DR: Efficient intron-mediated site-specific gene replacement and insertion approaches that generate mutations using the non-homologous end joining (NHEJ) pathway using the clustered regularly interspaced short palindromic repeats (CRISPR)– CRISPR-associated protein 9 (Cas9) system are reported.
Journal ArticleDOI
CRISPR/Cas9-mediated efficient and heritable targeted mutagenesis in tomato plants in the first and later generations.
TL;DR: It is demonstrated that the CRISPR/Cas9 system is an efficient tool for generating stable and heritable modifications in tomato plants through an Agrobacteria tumefaciens-mediated transformation method.
Journal ArticleDOI
Efficient generation of knock-in transgenic zebrafish carrying reporter/driver genes by CRISPR/Cas9-mediated genome engineering
TL;DR: It is proposed that CRISPR/Cas9-mediated knock-in will become a standard method for the generation transgenic zebrafish and be applied for the targeting integration of foreign genes into endogenous genomic loci.
Journal ArticleDOI
Rational design of a split-Cas9 enzyme complex.
Addison V. Wright,Samuel H. Sternberg,David W. Taylor,Brett T. Staahl,Jorge A Bardales,Jack E. Kornfeld,Jennifer A. Doudna +6 more
TL;DR: This study explores the structural features that enable Cas9 to bind and cleave target DNAs, and the results suggest a way of regulating Cas9 by physical separation of the catalytic domains from the rest of the protein scaffold, and proposes that split-Cas9 can act as a highly regulatable platform for genome-engineering applications.
Patent
CRISPR-Cas Nickase Systems, Methods And Compositions For Sequence Manipulation in Eukaryotes
Feng Zhang,Le Cong,Fei Ran +2 more
TL;DR: In this paper, the authors provide a system, methods, and compositions for manipulation of sequences and/or activities of target sequences, including vectors and vector systems, some of which encode one or more components of a CRISPR complex, and methods for the design and use of such vectors.
References
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Book
Molecular Cloning: A Laboratory Manual
TL;DR: Molecular Cloning has served as the foundation of technical expertise in labs worldwide for 30 years as mentioned in this paper and has been so popular, or so influential, that no other manual has been more widely used and influential.
疟原虫var基因转换速率变化导致抗原变异[英]/Paul H, Robert P, Christodoulou Z, et al//Proc Natl Acad Sci U S A
TL;DR: PfPMP1)与感染红细胞、树突状组胞以及胎盘的单个或多个受体作用,在黏附及免疫逃避中起关键的作�ly.
Journal ArticleDOI
Conserved seed pairing, often flanked by adenosines, indicates that thousands of human genes are microRNA targets
TL;DR: In a four-genome analysis of 3' UTRs, approximately 13,000 regulatory relationships were detected above the estimate of false-positive predictions, thereby implicating as miRNA targets more than 5300 human genes, which represented 30% of the gene set.
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.
Journal ArticleDOI
CRISPR RNA maturation by trans -encoded small RNA and host factor RNase III
Elitza Deltcheva,Krzysztof Chylinski,Krzysztof Chylinski,Cynthia M. Sharma,Karine Gonzales,Yanjie Chao,Zaid Ahmed Pirzada,Maria R. Eckert,Jörg Vogel,Emmanuelle Charpentier,Emmanuelle Charpentier +10 more
TL;DR: In this article, tracrRNA, a trans-encoded small RNA with 24-nucleotide complementarity to the repeat regions of crRNA precursor transcripts, is shown to direct the maturation of crRNAs by the activities of the widely conserved endogenous RNase III and the CRISPR-associated Csn1 protein.
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