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A Programmable Dual-RNA–Guided DNA Endonuclease in Adaptive Bacterial Immunity
- M. Jinek, Krzysztof Chylinski, Ines Fonfara, Michael H. Hauer, J. Doudna, E. Charpentier
- 17 August 2012
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.
CRISPR RNA maturation by trans-encoded small RNA and host factor RNase III
CRISPR/Cas systems constitute a widespread class of immunity systems that protect bacteria and archaea against phages and plasmids, and commonly use repeat/spacer-derived short crRNAs to silence…
Phylogeny of Cas9 determines functional exchangeability of dual-RNA and Cas9 among orthologous type II CRISPR-Cas systems
The reported collection of dual-RNA:Cas9 with associated PAMs expands the possibilities for multiplex genome editing and could provide means to improve the specificity of the RNA-programmable Cas9 tool.
The tracrRNA and Cas9 families of type II CRISPR-Cas immunity systems
CRISPR-Cas is a rapidly evolving RNA-mediated adaptive immune system that protects bacteria and archaea against mobile genetic elements. The system relies on the activity of short mature CRISPR RNAs…
Classification and evolution of type II CRISPR-Cas systems
- Krzysztof Chylinski, K. Makarova, E. Charpentier, E. Koonin
- BiologyNucleic acids research
- 11 April 2014
Phylogenomic analysis suggests that at least three cas genes, cas1, cas2 and cas4, and theCRISPR repeats of the type II-B system were acquired via recombination with a type I CRISPR-Cas locus, suggesting that type II CRISpr-Cas evolved via recombinations of mobile nuclease genes with type I loci.
CRISPR-Switch regulates sgRNA activity by Cre recombination for sequential editing of two loci
A precisely controlled sgRNA expression cassette that can be combined with widely-used Cre systems, termed CRISPR-Switch (SgRNA With Induction/Termination by Cre Homologous recombination), which substantially increases the versatility of gene editing through precise and rapid switching ON or OFF sg RNA activity, as well as switching OVER to secondary sgRNAs.
RNA sequencing uncovers antisense RNAs and novel small RNAs in Streptococcus pyogenes
- Anaïs Le Rhun, Yan Yan Beer, Johan Reimegård, Krzysztof Chylinski, E. Charpentier
- BiologyRNA biology
- 18 November 2015
It is shown that the expression profile of 9 sRNAs including 2 predicted regulatory elements is affected by the endoribonucleases RNase III and/or RNase Y, highlighting the critical role of these enzymes in sRNA regulation.
Methods and compositions for RNA-directed target DNA modification and for RNA-directed transcription modulation
A method for modifying a target DNA, the method comprising contacting the target DNA with a complex comprising a Cas9 polypeptide and an RNA directed to single molecule DNA, wherein said modification is cleavage of the targetDNA.
Bridge helix arginines play a critical role in Cas9 sensitivity to mismatches
It is demonstrated that arginines in the Cas9 bridge helix influence guide RNA, and target DNA binding and cleavage, and that R63 and R66 reduce Cas9 specificity by stabilizing the R-loop in the presence of mismatches.