Recent advances in the CRISPR genome editing tool set

Moon, Su Bin; Kim, Do Yon; Ko, Jeong Heon; Kim, Yong Sam

doi:10.1038/s12276-019-0339-7

Cited by 151 publications

(98 citation statements)

References 104 publications

(128 reference statements)

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“…deaminases have been generated (Moon et al, 2019). In the context of BIG-TREE, we employed AncBE4max, which displays a relatively high editing efficiency with low offtarget activity (Koblan et al, 2018).…”

Section: Discussionmentioning

confidence: 99%

“…In fact, base editors have shown a lower rate of indel formation and fewer off-target editing events than Cas9 (Komor et al, 2018). Over the past several years, various additional base editors have been engineered with different deaminases, targeting windows, editing efficiencies, and PAM specificities (Moon et al, 2019;Rees and Liu, 2018). We recently reported the development of a method called transient reporter for editing enrichment (TREE), which allows for bulk enrichment of base-edited cell populations, including hPSCs (Standage-Beier et al, 2019).…”

Section: Introductionmentioning

confidence: 99%

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BIG-TREE: Base-Edited Isogenic hPSC Line Generation Using a Transient Reporter for Editing Enrichment

et al. 2020

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Current CRISPR-targeted single-nucleotide modifications and subsequent isogenic cell line generation in human pluripotent stem cells (hPSCs) require the introduction of deleterious double-stranded DNA breaks followed by inefficient homology-directed repair (HDR). Here, we utilize Cas9 deaminase base-editing technologies to co-target genomic loci and an episomal reporter to enable single-nucleotide genomic changes in hPSCs without HDR. Together, this method entitled base-edited isogenic hPSC line generation using a transient reporter for editing enrichment (BIG-TREE) allows for single-nucleotide editing efficiencies of >80% across multiple hPSC lines. In addition, we show that BIG-TREE allows for efficient generation of loss-of-function hPSC lines via introduction of premature stop codons. Finally, we use BIG-TREE to achieve efficient multiplex editing of hPSCs at several independent loci. This easily adoptable method will allow for the precise and efficient base editing of hPSCs for use in developmental biology, disease modeling, drug screening, and cell-based therapies.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

BIG-TREE: Base-Edited Isogenic hPSC Line Generation Using a Transient Reporter for Editing Enrichment

et al. 2020

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“…The “DaZao” and transgenic Cas9 strain of B. mori were maintained in our laboratory[24]. Silkworm larvae were fed on fresh mulberry leaves and maintained at 25°C under standard conditions.…”

Section: Methodsmentioning

confidence: 99%

CRISPR/Cpf1 mediated genome editing enhancesBombyx moriresistance to BmNPV

Dong

et al. 2020

Preprint

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18CRISPR/Cas12a (Cpf1) is a single RNA-guided endonuclease that provides new 19 opportunities for targeted genome engineering through the CRISPR/Cas9 system. Only 20 AsCpf1 have been developed for insect genome editing, and the novel Cas12a orthologs 21 nucleases and editing efficiency require more study in insect. We compared three 22 Cas12a orthologs nucleases, AsCpf1, FnCpf1, and LbCpf1, for their editing efficiencies 23 and antiviral abilities in vitro. The three Cpf1 efficiently edited the BmNPV genome 24 and inhibited BmNPV replication in BmN-SWU1 cells. The antiviral ability of the 25 FnCpf1 system was more efficient than the SpCas9 system after infection by BmNPV. 26 We created FnCpf1×gIE1 and SpCas9×sgIE1 transgenic hybrid lines and evaluated the 27 gene editing efficiency of different systems at the same target site. We improved the 28 antiviral ability using the FnCpf1 system in transgenic silkworm. This study 29 demonstrated use of the CRISPR/Cpf1 system to achieve high editing efficiencies in 30 the silkworm, and illustrates the use of this technology for increasing disease resistance. 31Genome editing is a powerful tool that has been widely used in gene function, gene 36 therapy, pest control, and disease-resistant engineering in most parts of pathogens 37 research. Since the establishment of CRISPR/Cas9, powerful strategies for antiviral 38 therapy of transgenic silkworm have emerged. Nevertheless, there is still room to 39 expand the scope of genome editing tool for further application to improve antiviral 40 research. Here, we demonstrate that three Cpf1 endonuclease can be used efficiency 41 editing BmNPV genome in vitro and in vivo for the first time. More importantly, this 42 Cpf1 system could improve the resistance of transgenic silkworms to BmNPV compare 43 with Cas9 system, and no significant cocoons difference was observed between 44 transgenic lines infected with BmNPV and control. These broaden the range of 45 application of CRISPR for novel genome editing methods in silkworm and also enable 46 sheds light on antiviral therapy.47 49 Genome editing introduces DNA mutations in the form of insertions, deletions or 50 base substitutions within selected DNA sequences [1]. Clustered regularly interspaced 51 short palindromic repeats (CRISPR) gene editing technology has been used in gene 52 function research, genetic improvement, modelling biology and gene therapy [2-5]. 53 Three effector proteins of class 2 type V CRISPR systems, the CRISPR/CRISPR-54 associated 12a (Cas12a, known as Cpf1) proteins of Lachnosperaceae bacterium 55 (LbCpf1), Francisella novicida (FnCpf1) and Acidaminoccocus sp. (AsCpf1), have 56 been shown to efficiently edit mammalian cell genomes with more efficient genome 57 editing than the widely used Streptococcus pyogenes Cas9 (SpCas9) [6-8]. However, 58 the CRISPR/Cpf1 system was rarely used for insect genome editing research and 59 antiviral therapy [9].60 The silk industry (Bombyx mori, B. mori), suffers great economic losses due to B. 61 mori nucleopolyhedr...

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“…The efficiency of the genome editing therapy depend on the specificity of the DNA cleavage together with the prevention of collateral damage to the rest of the genome. The CRISPR/Cas9 system was proven as a suitable tool for stable and efficient genome editing as well as for high-throughput screening of mutations involved in oncogenesis and tumor progression [112][113][114][115]. The mostly used CRISPR/Cas9 system is the CRISPR system of Streptoccocus pyogenes (SpCas9), that recognize the short sequence 5 -NGG, where N represents any nucleotide and G represents guanine, and Cas9 is an nuclease guided by a single guide RNA (sgRNA) mediated by paring to the target sequence (reviewed in [116]).…”

Section: Genome Editingmentioning

confidence: 99%

Gene Therapy in Cancer Treatment: Why Go Nano?

et al. 2020

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The proposal of gene therapy to tackle cancer development has been instrumental for the development of novel approaches and strategies to fight this disease, but the efficacy of the proposed strategies has still fallen short of delivering the full potential of gene therapy in the clinic. Despite the plethora of gene modulation approaches, e.g., gene silencing, antisense therapy, RNA interference, gene and genome editing, finding a way to efficiently deliver these effectors to the desired cell and tissue has been a challenge. Nanomedicine has put forward several innovative platforms to overcome this obstacle. Most of these platforms rely on the application of nanoscale structures, with particular focus on nanoparticles. Herein, we review the current trends on the use of nanoparticles designed for cancer gene therapy, including inorganic, organic, or biological (e.g., exosomes) variants, in clinical development and their progress towards clinical applications.

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Recent advances in the CRISPR genome editing tool set

Cited by 151 publications

References 104 publications

BIG-TREE: Base-Edited Isogenic hPSC Line Generation Using a Transient Reporter for Editing Enrichment

BIG-TREE: Base-Edited Isogenic hPSC Line Generation Using a Transient Reporter for Editing Enrichment

CRISPR/Cpf1 mediated genome editing enhancesBombyx moriresistance to BmNPV

Gene Therapy in Cancer Treatment: Why Go Nano?

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