CRISPR Cpf1 proteins: structure, function and implications for genome editing

Safari, Fatemeh; Zare, Khadijeh; Negahdaripour, Manica; Barekati-Mowahed, Mazyar; Ghasemi, Younes

doi:10.1186/s13578-019-0298-7

Cited by 154 publications

(99 citation statements)

References 160 publications

(240 reference statements)

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“…It has been considered more effective than HDR‐mediated base‐pair substitution . To date, several base editing systems, for instance BE3, BE4, Targeted‐AID, and dCpf1‐BE have been used in various organisms including major crops. These systems utilize Cas9 or Cpf1 systems for recruiting cytidine deaminases, which generate specific C–T alterations by using DNA mismatch repair pathways …”

Section: E‐crisp/cas Systemsmentioning

confidence: 99%

CRISPR/Cas Systems in Genome Editing: Methodologies and Tools for sgRNA Design, Off‐Target Evaluation, and Strategies to Mitigate Off‐Target Effects

et al. 2020

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Life sciences have been revolutionized by genome editing (GE) tools, including zinc finger nucleases, transcription activator‐Like effector nucleases, and CRISPR (clustered regulatory interspaced short palindromic repeats)/Cas (CRISPR‐associated) systems, which make the targeted modification of genomic DNA of all organisms possible. CRISPR/Cas systems are being widely used because of their accuracy, efficiency, and cost‐effectiveness. Various classes of CRISPR/Cas systems have been developed, but their extensive use may be hindered by off‐target effects. Efforts are being made to reduce the off‐target effects of CRISPR/Cas9 by generating various CRISPR/Cas systems with high fidelity and accuracy. Several approaches have been applied to detect and evaluate the off‐target effects. Here, the current GE tools, the off‐target effects generated by GE technology, types of off‐target effects, mechanisms of off‐target effects, major concerns, and outcomes of off‐target effects in plants and animals are summarized. The methods to detect off‐target effects, tools for single‐guide RNA (sgRNA) design, evaluation and prediction of off‐target effects, and strategies to increase the on‐target efficiency and mitigate the off‐target impact on intended genome‐editing outcomes are summarized.

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Section: E‐crisp/cas Systemsmentioning

confidence: 99%

CRISPR/Cas Systems in Genome Editing: Methodologies and Tools for sgRNA Design, Off‐Target Evaluation, and Strategies to Mitigate Off‐Target Effects

et al. 2020

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“…In this study, one important application of DocMF was to quickly and accurately identify the PAMs of any Crispr-Cas system (Figure 3-4). The Crispr-Cas system was initially identified in the prokaryotic immune system and was quickly adopted as a reliable genome engineering tool (24) (31). The PAM sequence is adjacent to the target site and is essential for Cas endonuclease specificity.…”

Section: Discussionmentioning

confidence: 99%

“…To increase the number of potential genome editing sites, we are in urgent need of new Cas (that recognize PAM sequences beyond the commonly used 5'-NGG-3' site for SpCas9 (16). DocMF could be a very useful tool for characterizing the novel Cas protein PAM sequences with the following advantages: 1) a universal system for different Crispr-Cas systems, with the same DNB pools PAMs, a single guide RNA, and the production of a staggered DNA double-stranded break (31). Our characterization of VeCas9 and BvCpf1 helps to expand the existing CRISPR toolbox and provide more promising candidates for genome engineering.…”

Section: Discussionmentioning

confidence: 99%

DNB-Based On-Chip Motif Finding (DocMF): a High-Throughput Method to Profile Different Types of Protein-DNA Interactions

Wang

et al. 2019

Preprint

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DNA-protein interactions play pivotal roles in many cellular activities, such as replication, development, and DNA repair. Here we report a DNB-based on-chip Motif Finding (DocMF) system that utilizes next-generation-sequencing chips to profile protein binding or cleaving activity toward several hundred millions or billions of DNA sequences. Using DocMF, we successfully identified a variety of endonuclease recognition sites and the protospacer-adjacent motif (PAM) sequences of different CRISPR effector proteins. Compared with the widely used PAM depletion assay, our DocMF platform can simultaneously screen both 5' and 3' sequences adjacent to the protospacer to identify PAMs with high coverage using the same DNB library pool. For the well-studied SpCas9, our DocMF platform not only identified its common PAMs, 5'-NGG-3' (~89.9%), but also revealed a small proportion of noncanonical 5'-NAG-3' (~5%) and 5'-NGA-3' (~1.6%) PAMs. We also used the DocMF to assay two uncharacterized Cas endonucleases, VeCas9 and BvCpf1. VeCas9 PAMs were not detected by the conventional PAM depletion method. However, DocMF successfully revealed that both VeCas9 and BvCpf1 required broader and more complicated PAM sequences for target DNA recognition. VeCas9 preferred the R-rich motifs, whereas BvCpf1 used the T-rich PAMs. Moreover, after slightly changing the experimental protocol and analysis threshold, we observed that dCas9, a DNA binding protein lacking endonuclease activity, preferably binds to the previously reported PAMs 5'-NGG-3'. In summary, our studies demonstrate that DocMF is the first tool with the capacity to exhaustively assay both the binding and the cutting site preferences of different DNA-binding proteins.

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“…Using the Cpf1 endonuclease derived from Acidaminococcus sp. BV3L6 and Lachnospiraceae ND2006 bacteria, it was found that the effect of this system on eukaryotic cells is compatible with the CRISPR-Cas9 [38]. However, functional Cpf1 does not require the tracrRNA, but crRNA is needed.…”

Section: Crispr-cpf1(cas12)mentioning

confidence: 95%

An overview of applications of CRISPR-Cas technologies in biomedical engineering

Jamehdor¹,

Kaboli

Naserian

et al. 2020

Folia Histochem Cytobiol.

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Clustered Regulatory Interspaced Short Palindromic Repeats (CRISPR) is one of the major genome editing systems and allows changing DNA levels of an organism. Among several CRISPR categories, the CRISPR-Cas9 system has shown a remarkable progression rate over its lifetime. Recently, other tools including CRISPR-Cas12 and CRISPR-Cas13 have been introduced. CRISPR-Cas9 system has played a key role in the industrial cell factory's production and improved our understanding of genome function. Additionally, this system has been used as one of the major genome editing systems for the diagnosis and treatment of several infectious and non-infectious diseases. In this review, we discuss CRISPR biology, its versatility, and its application in biomedical engineering.

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CRISPR Cpf1 proteins: structure, function and implications for genome editing

Cited by 154 publications

References 160 publications

CRISPR/Cas Systems in Genome Editing: Methodologies and Tools for sgRNA Design, Off‐Target Evaluation, and Strategies to Mitigate Off‐Target Effects

CRISPR/Cas Systems in Genome Editing: Methodologies and Tools for sgRNA Design, Off‐Target Evaluation, and Strategies to Mitigate Off‐Target Effects

DNB-Based On-Chip Motif Finding (DocMF): a High-Throughput Method to Profile Different Types of Protein-DNA Interactions

An overview of applications of CRISPR-Cas technologies in biomedical engineering

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