Precise Editing of a Target Base in the Rice Genome Using a Modified CRISPR/Cas9 System

Lu, Yuming; Zhu, Jian‐Kang

doi:10.1016/j.molp.2016.11.013

Cited by 367 publications

(200 citation statements)

References 12 publications

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“…Recently, CRISPR/Cas9 machinery has been modified to induce a genetic change without introducing a doublestranded break, called Bbase editing.^This method utilizes cytidine deaminases and guide RNA to convert cytidine to uridine, resulting in a change to thymidine [15]. This technology has been used to efficiently introduce genetic changes into a variety of species, including plant, yeast, sea urchin, mouse zygotes, and human cells and tripronuclear zygotes [15][16][17][18][19][20][21][22][23][24][25][26]. While this technology still needs to be refined as mosaicism is still observed, it provides a new method that may introduce genetic changes more efficiently without having to damage DNA.…”

Section: Mybpc3mentioning

confidence: 99%

Editing the human genome: where ART and science intersect

Hershlag

Bristow

2018

J Assist Reprod Genet

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The rapid development of gene-editing technologies has led to an exponential rise in both basic and translational research initiatives studying molecular processes and investigating possible clinical applications. Early experiments using genome editing to study human embryo development have contradicted findings in studies on model organisms. Additionally, a series of four experiments over the past 2 years set out to investigate the possibilities of introducing genetic modifications to human embryos, each with varying levels of success. Here, we discuss the key findings of these studies, including the efficiency, the safety, the potential untoward effects, major flaws of the studies, and emerging alternative genome editing methods that may allow overcoming the hurdles encountered so far. Given these results, we also raise several questions about the clinical utilization of germline gene editing: For which indications is gene editing appropriate? How do gene-editing technologies compare with genetic testing methods currently used for screening embryos? What are the ethical considerations we should be concerned about? While further research is underway, and our understanding of how to implement this technology continues to evolve, it is critical to contemplate if and how it should be translated from the bench to clinical practice.

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Section: Mybpc3mentioning

confidence: 99%

Editing the human genome: where ART and science intersect

Hershlag

Bristow

2018

J Assist Reprod Genet

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“…For better efficiency, the cytidine deaminase can be fused to a nickase that cleaves the non-edited strand and be associated to the uracil glycosylase inhibitor (UGI) that inhibits base-excision repair, to limit indel formation at the cleavage site [46]. This approach was successfully used to edit endogenous genes in tomato, rice, maize and wheat with the Cas9 D10A at frequencies of up to 43.48% [47][48][49][50], which is so far much more efficient than HDR-mediated gene replacement. However, the base-editing efficiency seems to be dependent on the target sequence and the criteria for the selection of adapted sgRNAs request further investigations [47].…”

Section: Spcas9: Nuclease Nickase and Dead Versionsmentioning

confidence: 99%

Towards mastering CRISPR-induced gene knock-in in plants: Survey of key features and focus on the model Physcomitrella patens

Collonnier

Guyon‐Debast

Maclot

et al. 2017

Methods

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a b s t r a c tBeyond its predominant role in human and animal therapy, the CRISPR-Cas9 system has also become an essential tool for plant research and plant breeding. Agronomic applications rely on the mastery of gene inactivation and gene modification. However, if the knock-out of genes by non-homologous end-joining (NHEJ)-mediated repair of the targeted double-strand breaks (DSBs) induced by the CRISPR-Cas9 system is rather well mastered, the knock-in of genes by homology-driven repair or end-joining remains difficult to perform efficiently in higher plants. In this review, we describe the different approaches that can be tested to improve the efficiency of CRISPR-induced gene modification in plants, which include the use of optimal transformation and regeneration protocols, the design of appropriate guide RNAs and donor templates and the choice of nucleases and means of delivery. We also present what can be done to orient DNA repair pathways in the target cells, and we show how the moss Physcomitrella patens can be used as a model plant to better understand what DNA repair mechanisms are involved, and how this knowledge could eventually be used to define more performant strategies of CRISPR-induced gene knock-in.

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“…Early demonstration in human cells by Komor et al (2016) yielded no detectable base editing at the known dCas9 off-target sites, and that, base editors in human cells do not induce untargeted C→T conversion throughout the genome. Later on, Lu and Zhu (2016) tested the applicability of this modified CRISPR/Cas9 system in the rice callus of Zhonghua11 (ZH11) using Agrobacterium-mediated transformation. In their study, they fused rat APOBEC1 to the N-terminus of Cas9(D10A) using the unstructured 16-residue peptide XTEN as linker, and constructed it into a binary vector containing maize ubiquitin promoter.…”

Section: Milestones Of Crispr/cas9 In Crop Biotechnologymentioning

confidence: 99%

“…Results showed similar base-editing results in human cells, wherein respective C→T substitution frequency in NRT1.1B and SLR1 was 1.4%-11.5%, while C→G replacement frequency was at 1.6%-3.9%. Lu and Zhu (2016) declared that the possible cause of the lower base-editing efficiency on NRT1.1B was the lower targeting efficiency of the gRNA for this gene. Furthermore, to demonstrate feasibility of this new approach in plant breeding program, Lu and Zhu (2016) generated stable transgenic seedlings from the hygromacin-resistant callus.…”

Section: Milestones Of Crispr/cas9 In Crop Biotechnologymentioning

confidence: 99%

“…Lu and Zhu (2016) declared that the possible cause of the lower base-editing efficiency on NRT1.1B was the lower targeting efficiency of the gRNA for this gene. Furthermore, to demonstrate feasibility of this new approach in plant breeding program, Lu and Zhu (2016) generated stable transgenic seedlings from the hygromacin-resistant callus. Sanger sequencing and RFLP results of SLR1 consistently confirmed the base replacement in the six transgenic lines which showed a semi-dwarf phenotype.…”

Section: Milestones Of Crispr/cas9 In Crop Biotechnologymentioning

confidence: 99%

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CRISPR/CAS9 as a Powerful Tool for Crop Improvement

et al. 2017

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This is an Open-Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. Recently, a modified version of CRISPR/Cas9 with added novel functions has been developed that enables programmable direct irreversible conversion of a target DNA base. In this review, we summarized the milestone applications of CRISPR/ Cas9 in plants with a focus on major crops. We also present the implications of an improved version of this technology in the current plant breeding programs.

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Precise Editing of a Target Base in the Rice Genome Using a Modified CRISPR/Cas9 System

Cited by 367 publications

References 12 publications

Editing the human genome: where ART and science intersect

Editing the human genome: where ART and science intersect

Towards mastering CRISPR-induced gene knock-in in plants: Survey of key features and focus on the model Physcomitrella patens

CRISPR/CAS9 as a Powerful Tool for Crop Improvement

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