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BigField GEG Tech's insight:
The authors discuss experimental considerations, limitations and critical aspects which will guide the investigator for successful implementation of the genome editing technology in human PSCs using designer nucleases. 
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BigField GEG Tech's insight:
Reports of the simple generation of animal models and genome engineering of cells raised questions about targeting precision. Conflicting early reports led the field to believe that CRISPR/Cas9 system was promiscuous, leading to a variety of strategies for improving specificity and increasingly sensitive methods to detect off-target events. However, other studies have suggested that CRISPR/Cas9 is a highly specific genome-editing tool. This review will focus on deciphering and interpreting these seemingly opposing claims.
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BigField GEG Tech's insight:
The authors provide a detailed protocol for design and construction of TALEN and CRISPR vectors, testing of their nuclease activity, and delivery of TALEN or CRISPR vectors into hPSCs. Moreover, they describe the identification of edited hPSC clones without antibiotic selection, including their clonal selection, genotyping, and expansion for downstream applications.
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BigField GEG Tech's insight:
In this study, The authors report the application of CRISPR/Cas9-mediated knock-in of a reporter cassette, which does not disrupt endogenous target genes in mouse haploid ESCs. In this way, they engineered the neural marker gene Sox1 locus and verified the precise insertion of the P2A-Venus reporter cassette and its functionality by monitoring neural differentiation.
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BigField GEG Tech's insight:
Off-target effects are a critical issue for genome editing and transcriptome modulation. Here the authors review the current status on the target specificity of the CRISPR-Cas9 system.
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BigField GEG Tech's insight:
CRISPR-Cas systems have immense biotechnological utility. A recent study reveals the potential of the Cpf1 nuclease to complement and extend the existing CRISPR-Cas9 genome-editing tools.
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BigField GEG Tech's insight:
The authors of this study previously identified he first examples of proteins that inhibit a CRISPR–Cas system. Here they performed biochemical and in vivo investigations of three of these anti-CRISPR proteins, and show that each inhibits CRISPR–Cas activity through a distinct mechanism. The diverse sequences and mechanisms of action of these anti-CRISPR proteins imply an independent evolution, and foreshadow the existence of other means by which proteins may alter CRISPR–Cas function.
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BigField GEG Tech's insight:
Here, the scientists showed that limited low-dose irradiation (LDI) using either γ-ray or x-ray exposure (0.4 Gy) significantly enhanced HR frequency, possibly through induction of DNA repair/recombination machinery including ataxia-telangiectasia mutated, histone H2A.X and RAD51 proteins. LDI could also increase HR efficiency by more than 30-fold when combined with the targeting tools zinc finger nucleases, transcription activator-like effector nucleases, and clustered regularly interspaced short palindromic repeats.
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BigField GEG Tech's insight:
The author review early experiments using meganucleases, zinc-finger nucleases (ZFN), and transcription-activator like effector nucleases (TALENs) to contract trinucleotide repeats, and discuss the possibility of using CRISPR-Cas nucleases to the same end. Although this is a nascent field, he explores the possibility of designing nucleases and effectively delivering them in the context of gene therapy.
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BigField GEG Tech's insight:
The authors review the fast developing technology of targeted genome engineering using site specific programmable nucleases zinc finger nucleases (ZFNs), transcription activator like nucleases (TALENs) and cluster regulatory interspaced short palindromic repeat/CRISPR associated proteins (CRISPR/Cas) based RNA-guided DNA endonucleases (RGENs) and their different characteristics including pros and cons of genome modifications by these nucleases. They have further discussed different types of delivery methods to induce gene editing, novel development in genetic engineering other than nucleases and future prospects.
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PLOS ONE: an inclusive, peer-reviewed, open-access resource from the PUBLIC LIBRARY OF SCIENCE. Reports of well-performed scientific studies from all disciplines freely available to the whole world.
BigField GEG Tech's insight:
Smart strategy which allows a very simple, efficient and fast assay for detecting induced mutations It will be surely widely used in the area of genome editing.
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The authors discuss experimental considerations, limitations and critical aspects which will guide the investigator for successful implementation of the genome editing technology in human PSCs using designer nucleases.