Kinetic analysis of Cas12a and Cas13a RNA-Guided nucleases for development of improved CRISPR-Based diagnostics
Eric Nalefski,Nidhi Patel,Philip J.Y. Leung,Zeba Islam,Remy M. Kooistra,Ishira Parikh,Estelle Marion,Gavin J. Knott,Gavin J. Knott,Jennifer A. Doudna,Anne-Laure M. Le Ny,Damian Madan +11 more
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TLDR
In this article, the authors examined the kinetics of single-turnover target and multi-turnoff trans-nuclease activities of both enzymes and showed that these enzymes are kinetically adapted to play distinct roles in bacterial adaptive immunity and show how kinetic analysis can be applied to CRISPR-based diagnostics.About:
This article is published in iScience.The article was published on 2021-08-18 and is currently open access. It has received 38 citations till now. The article focuses on the topics: Trans-activating crRNA & CRISPR.read more
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Multiplexed CRISPR-based microfluidic platform for clinical testing of respiratory viruses and identification of SARS-CoV-2 variants
Nicole L. Welch,M. Zhu,Catherine Hua,Juliane Weller,Marzieh Ezzaty Mirhashemi,Tien G. Nguyen,Sreekar Mantena,Matthew R. Bauer,Bennett M. Shaw,Cheri M. Ackerman,Sri Gowtham Thakku,Megan Tse,Jared Kehe,Marie-Martine Uwera,Jacqueline S. Eversley,D.A. Bielwaski,Graham McGrath,Joseph Braidt,Jeremy Johnson,Felecia Cerrato,Gage K. Moreno,Lydia A. Krasilnikova,Brittany A. Petros,Gabrielle Gionet,Ewa King,Richard C. Huard,S. Jalbert,Michael L. Cleary,N. Fitzgerald,Stacey Gabriel,Glen R. Gallagher,Sandra Smole,Lawrence C. Madoff,Catherine M. Brown,Matthew W. Keller,Malania M. Wilson,M. Kirby,John R. Barnes,Daniel J. Park,Katherine J. Siddle,Christian T. Happi,Deborah T. Hung,Michael Springer,Bronwyn MacInnis,Jacob E. Lemieux,Eric S. Rosenberg,John A. Branda,Paul C. Blainey,Pardis C. Sabeti,Cameron Myhrvold +49 more
TL;DR: In this article , a cost-effective virus and variant detection platform, called microfluidic Combinatorial Arrayed Reactions for Multiplexed Evaluation of Nucleic acids (mCARMEN), is presented.
Journal ArticleDOI
CRISPR/Cas-powered nanobiosensors for diagnostics
TL;DR: In this article , the authors explore some of the most recent advances in the field of CRISPR-powered nanotechnological biosensors, which offer tremendous potential for next-generation diagnostics of multiple targets, especially at the point of care and without any target amplification.
Journal ArticleDOI
CRISPR/Cas-powered nanobiosensors for diagnostics.
Quynh Anh Phan,Ebrahim Mostafavi,Linh B. Truong,David Medina-Cruz,Can Dincer,Ebrahim Mostafavi +5 more
TL;DR: In this article, the authors explore some of the most recent advances in the field of CRISPR-powered nanotechnological biosensors, which offer tremendous potential for next-generation diagnostics of multiple targets, especially at the point of care and without any target amplification.
Journal ArticleDOI
Electrochemical biosensor for nucleic acid amplification-free and sensitive detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA via CRISPR/Cas13a trans-cleavage reaction
TL;DR: In this article , an electrochemical biosensor based on the trans-cleavage activity of CRISPR/Cas13a was developed for rapid, sensitive, and nucleic acid-amplification-free detection of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).
Journal ArticleDOI
Enzyme Kinetics and Detector Sensitivity Determine Limits of Detection of Amplification-Free CRISPR-Cas12 and CRISPR-Cas13 Diagnostics.
Diego A. Huyke,Ashwin Ramachandran,Vladimir I. Bashkirov,Efthalia K. Kotseroglou,Theofilos Kotseroglou,Juan G. Santiago +5 more
TL;DR: The results suggest that successful detection of target requires cleavage of at least 0.1% of the fluorescent reporter molecules, and demonstrate the fundamental nature by which kinetic rates and background signal limit LoDs and thus highlight areas of improvement for the emerging field of CRISPR diagnostics.
References
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Journal ArticleDOI
Cpf1 is a single RNA-guided endonuclease of a class 2 CRISPR-Cas system.
Bernd Zetsche,Jonathan S. Gootenberg,Omar O. Abudayyeh,Ian Slaymaker,Kira S. Makarova,Patrick Essletzbichler,Sara E. Volz,Julia Joung,John van der Oost,Aviv Regev,Aviv Regev,Eugene V. Koonin,Feng Zhang +12 more
TL;DR: In this paper, the authors characterized Cpf1, a putative class 2 CRISPR effector, which is a single RNA-guided endonuclease lacking tracrRNA and utilizes a T-rich protospacer-adjacent motif.
Journal ArticleDOI
CRISPR-Cas12a target binding unleashes indiscriminate single-stranded DNase activity
Janice S. Chen,Enbo Ma,Lucas B. Harrington,Maria Da Costa,Xinran Tian,Joel M. Palefsky,Jennifer A. Doudna +6 more
TL;DR: It is shown that RNA-guided DNA binding unleashes indiscriminate single-stranded DNA cleavage activity by Cas12a that completely degrades ssDNA molecules, which is also a property of other type V CRISPR-Cas12 enzymes.
Journal ArticleDOI
DNA interrogation by the CRISPR RNA-guided endonuclease Cas9
TL;DR: It is shown that both binding and cleavage of DNA by Cas9–RNA require recognition of a short trinucleotide protospacer adjacent motif (PAM) and that PAM interactions trigger Cas9 catalytic activity.
Journal ArticleDOI
C2c2 is a single-component programmable RNA-guided RNA-targeting CRISPR effector
Omar O. Abudayyeh,Jonathan S. Gootenberg,Silvana Konermann,Silvana Konermann,Silvana Konermann,Julia Joung,Julia Joung,Julia Joung,Ian Slaymaker,Ian Slaymaker,Ian Slaymaker,David Benjamin Turitz Cox,Sergey Shmakov,Sergey Shmakov,Kira S. Makarova,Ekaterina Semenova,Leonid Minakhin,Konstantin Severinov,Konstantin Severinov,Konstantin Severinov,Aviv Regev,Aviv Regev,Eric S. Lander,Eric S. Lander,Eric S. Lander,Eugene V. Koonin,Feng Zhang +26 more
TL;DR: LshC2c2 is a RNA-guided RNase which requires the activity of its two HEPN domains, suggesting previously unidentified mechanisms of RNA targeting and degradation by CRISPR systems.
Journal ArticleDOI
Discovery and Functional Characterization of Diverse Class 2 CRISPR-Cas Systems
Sergey Shmakov,Omar O. Abudayyeh,Kira S. Makarova,Yuri I. Wolf,Jonathan S. Gootenberg,Ekaterina Semenova,Leonid Minakhin,Julia Joung,Silvana Konermann,Konstantin Severinov,Konstantin Severinov,Konstantin Severinov,Feng Zhang,Eugene V. Koonin +13 more
TL;DR: In this article, three distinct Class 2 CRISPR-Cas systems are described, two of which contain RuvC-like endonuclease domains distantly related to Cpf1.
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