Targeted mutagenesis in <i>Arabidopsis thaliana</i> using CRISPR‐Cas12b/C2c1

Wu, Fan; Qiao, Xinyu; Zhao, Yafei; Zhang, Ziyi; Gao, Yifan; Shi, Ling‐Feng; Du, Haokun; Wang, Lulu; Zhang, Yajie; Zhang, Yu; Liu, Langyu; Wang, Quan; Kong, Delong

doi:10.1111/jipb.12944

Cited by 26 publications

(9 citation statements)

References 15 publications

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“…Cas12b (formerly C2c1) is a class 2 type V-B endonuclease and represents the third major CRISPR system that has been used for plant genome editing ( Shmakov et al, 2015 ; Ming et al, 2020 ; Wang et al, 2020d ; Wu et al, 2020 ). Like Cas12a, Cas12b recognizes T-rich PAM sequences, TTN, and generates staggered DSBs (7-nt 5′-overhangs) 23 bp upstream of the PAM on the non-target strand ( Shmakov et al, 2015 ; Liu et al, 2017b ).…”

Section: Crispr/cas12b Systemmentioning

confidence: 99%

Molecular and Computational Strategies to Increase the Efficiency of CRISPR-Based Techniques

et al. 2022

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The prokaryote-derived Clustered Regularly Interspaced Palindromic Repeats (CRISPR)/Cas mediated gene editing tools have revolutionized our ability to precisely manipulate specific genome sequences in plants and animals. The simplicity, precision, affordability, and robustness of this technology have allowed a myriad of genomes from a diverse group of plant species to be successfully edited. Even though CRISPR/Cas, base editing, and prime editing technologies have been rapidly adopted and implemented in plants, their editing efficiency rate and specificity varies greatly. In this review, we provide a critical overview of the recent advances in CRISPR/Cas9-derived technologies and their implications on enhancing editing efficiency. We highlight the major efforts of engineering Cas9, Cas12a, Cas12b, and Cas12f proteins aiming to improve their efficiencies. We also provide a perspective on the global future of agriculturally based products using DNA-free CRISPR/Cas techniques. The improvement of CRISPR-based technologies efficiency will enable the implementation of genome editing tools in a variety of crop plants, as well as accelerate progress in basic research and molecular breeding.

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Section: Crispr/cas12b Systemmentioning

confidence: 99%

Molecular and Computational Strategies to Increase the Efficiency of CRISPR-Based Techniques

et al. 2022

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“…Moreover, it is advisable to test multiple nuclease orthologs to identify the most efficient variants for plant genome editing. An examination of several Cas12b orthologs revealed that AaCas12b was the most efficient one in rice (Ming et al 2020), whereas other variants, AacCas12b, BvCas12 and BhCas12b v4, were demonstrated to work well in rice, cotton and Arabidopsis (Ming et al 2020;Wang et al 2020d;Wu et al 2020). Another example of mixed results is ScCas9, which was originally reported to have a minimal PAM requirement of NNG (Chatterjee et al 2018).…”

Section: Chromosomal Rearrangementmentioning

confidence: 99%

Novel CRISPR/Cas applications in plants: from prime editing to chromosome engineering

Huang

Puchta

2021

Transgenic Res

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In the last years, tremendous progress has been made in the development of CRISPR/Cas-mediated genome editing tools. A number of natural CRISPR/Cas nuclease variants have been characterized. Engineered Cas proteins have been developed to minimize PAM restrictions, off-side effects and temperature sensitivity. Both kinds of enzymes have, by now, been applied widely and efficiently in many plant species to generate either single or multiple mutations at the desired loci by multiplexing. In addition to DSB-induced mutagenesis, specifically designed CRISPR/Cas systems allow more precise gene editing, resulting not only in random mutations but also in predefined changes. Applications in plants include gene targeting by homologous recombination, base editing and, more recently, prime editing. We will evaluate these different technologies for their prospects and practical applicability in plants. In addition, we will discuss a novel application of the Cas9 nuclease in plants, enabling the induction of heritable chromosomal rearrangements, such as inversions and translocations. This technique will make it possible to change genetic linkages in a programmed way and add another level of genome engineering to the toolbox of plant breeding. Also, strategies for tissue culture free genome editing were developed, which might be helpful to overcome the transformation bottlenecks in many crops. All in all, the recent advances of CRISPR/Cas technology will help agriculture to address the challenges of the twenty-first century related to global warming, pollution and the resulting food shortage.

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“…Furthermore, Cas12b induces staggered cuts distal from the PAM sequence resulting in 6-8 nt long 5'-overhangs (Teng et al, 2018). Most recently, the CRISPR/Cas12b system was engineered for gene editing in plants and was efficiently applied in both, mono and dicotyledons (Ming et al, 2020;Wu et al, 2020).…”

Section: Crispr/cas Systemsmentioning

confidence: 99%