scholarly journals Plant genome editing made easy: targeted mutagenesis in model and crop plants using the CRISPR/Cas system

Plant Methods ◽  
2013 ◽  
Vol 9 (1) ◽  
pp. 39 ◽  
Author(s):  
Khaoula Belhaj ◽  
Angela Chaparro-Garcia ◽  
Sophien Kamoun ◽  
Vladimir Nekrasov
2019 ◽  
Vol 20 (16) ◽  
pp. 4045 ◽  
Author(s):  
Ali Razzaq ◽  
Fozia Saleem ◽  
Mehak Kanwal ◽  
Ghulam Mustafa ◽  
Sumaira Yousaf ◽  
...  

Increasing agricultural productivity via modern breeding strategies is of prime interest to attain global food security. An array of biotic and abiotic stressors affect productivity as well as the quality of crop plants, and it is a primary need to develop crops with improved adaptability, high productivity, and resilience against these biotic/abiotic stressors. Conventional approaches to genetic engineering involve tedious procedures. State-of-the-art OMICS approaches reinforced with next-generation sequencing and the latest developments in genome editing tools have paved the way for targeted mutagenesis, opening new horizons for precise genome engineering. Various genome editing tools such as transcription activator-like effector nucleases (TALENs), zinc-finger nucleases (ZFNs), and meganucleases (MNs) have enabled plant scientists to manipulate desired genes in crop plants. However, these approaches are expensive and laborious involving complex procedures for successful editing. Conversely, CRISPR/Cas9 is an entrancing, easy-to-design, cost-effective, and versatile tool for precise and efficient plant genome editing. In recent years, the CRISPR/Cas9 system has emerged as a powerful tool for targeted mutagenesis, including single base substitution, multiplex gene editing, gene knockouts, and regulation of gene transcription in plants. Thus, CRISPR/Cas9-based genome editing has demonstrated great potential for crop improvement but regulation of genome-edited crops is still in its infancy. Here, we extensively reviewed the availability of CRISPR/Cas9 genome editing tools for plant biotechnologists to target desired genes and its vast applications in crop breeding research.


2020 ◽  
Vol 3 (1) ◽  
Author(s):  
Keishi Osakabe ◽  
Naoki Wada ◽  
Tomoko Miyaji ◽  
Emi Murakami ◽  
Kazuya Marui ◽  
...  

Abstract Genome editing in plants has advanced greatly by applying the clustered regularly interspaced short palindromic repeats (CRISPRs)-Cas system, especially CRISPR-Cas9. However, CRISPR type I—the most abundant CRISPR system in bacteria—has not been exploited for plant genome modification. In type I CRISPR-Cas systems, e.g., type I-E, Cas3 nucleases degrade the target DNA in mammals. Here, we present a type I-D (TiD) CRISPR-Cas genome editing system in plants. TiD lacks the Cas3 nuclease domain; instead, Cas10d is the functional nuclease in vivo. TiD was active in targeted mutagenesis of tomato genomic DNA. The mutations generated by TiD differed from those of CRISPR/Cas9; both bi-directional long-range deletions and short indels mutations were detected in tomato cells. Furthermore, TiD can be used to efficiently generate bi-allelic mutant plants in the first generation. These findings indicate that TiD is a unique CRISPR system that can be used for genome engineering in plants.


2019 ◽  
Vol 20 (5) ◽  
pp. 1155 ◽  
Author(s):  
Guillaume Bernard ◽  
David Gagneul ◽  
Harmony Alves Dos Santos ◽  
Audrey Etienne ◽  
Jean-Louis Hilbert ◽  
...  

CRISPR/Cas9 (Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR associated with protein CAS9) is a genome-editing tool that has been extensively used in the last five years because of its novelty, affordability, and feasibility. This technology has been developed in many plant species for gene function analysis and crop improvement but has never been used in chicory (Cichorium intybus L.). In this study, we successfully applied CRISPR/Cas9-mediated targeted mutagenesis to chicory using Agrobacterium rhizogenes-mediated transformation and protoplast transfection methods. A U6 promoter (CiU6-1p) among eight predicted U6 promoters in chicory was selected to drive sgRNA expression. A binary vector designed to induce targeted mutations in the fifth exon of the chicory phytoene desaturase gene (CiPDS) was then constructed and used to transform chicory. The mutation frequency was 4.5% with the protoplast transient expression system and 31.25% with A. rhizogenes-mediated stable transformation. Biallelic mutations were detected in all the mutant plants. The use of A. rhizogenes-mediated transformation seems preferable as the regeneration of plants is faster and the mutation frequency was shown to be higher. With both transformation methods, foreign DNA was integrated in the plant genome. Hence, selection of vector (transgene)-free segregants is required. Our results showed that genome editing with CRISPR/Cas9 system can be efficiently used with chicory, which should facilitate and accelerate genetic improvement and functional biology.


2018 ◽  
Vol 131 ◽  
pp. 31-36 ◽  
Author(s):  
Mostafa Abdelrahman ◽  
Abdullah M. Al-Sadi ◽  
Alireza Pour-Aboughadareh ◽  
David J. Burritt ◽  
Lam-Son Phan Tran

2018 ◽  
pp. 47-54 ◽  
Author(s):  
Haris Khurshid ◽  
Sohail Ahmad Jan ◽  
Zabta Khan Shinwari ◽  
Muhammad Jamal ◽  
Sabir Hussain Shah
Keyword(s):  

2021 ◽  
Vol 22 (14) ◽  
pp. 7456
Author(s):  
Mousa A. Alghuthaymi ◽  
Aftab Ahmad ◽  
Zulqurnain Khan ◽  
Sultan Habibullah Khan ◽  
Farah K. Ahmed ◽  
...  

Rapid developments in the field of plant genome editing using clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein (Cas) systems necessitate more detailed consideration of the delivery of the CRISPR system into plants. Successful and safe editing of plant genomes is partly based on efficient delivery of the CRISPR system. Along with the use of plasmids and viral vectors as cargo material for genome editing, non-viral vectors have also been considered for delivery purposes. These non-viral vectors can be made of a variety of materials, including inorganic nanoparticles, carbon nanotubes, liposomes, and protein- and peptide-based nanoparticles, as well as nanoscale polymeric materials. They have a decreased immune response, an advantage over viral vectors, and offer additional flexibility in their design, allowing them to be functionalized and targeted to specific sites in a biological system with low cytotoxicity. This review is dedicated to describing the delivery methods of CRISPR system into plants with emphasis on the use of non-viral vectors.


2021 ◽  
pp. 096366252199979
Author(s):  
Robert D.J. Smith ◽  
Sarah Hartley ◽  
Patrick Middleton ◽  
Tracey Jewitt

Citizen and stakeholder engagement is frequently portrayed as vital for socially accountable science policy but there is a growing understanding of how institutional dynamics shape engagement exercises in ways that prevent them from realising their full potential. Limited attention has been devoted to developing the means to expose institutional features, allow policy-makers to reflect on how they will shape engagement and respond appropriately. Here, therefore, we develop and test a methodological framework to facilitate pre-engagement institutional reflexivity with one of the United Kingdom’s eminent science organisations as it grappled with a new, high-profile and politicised technology, genome editing. We show how this approach allowed policy-makers to reflect on their institutional position and enrich decision-making at a time when they faced pressure to legitimate decisions with engagement. Further descriptions of such pre-engagement institutional reflexivity are needed to better bridge theory and practice in the social studies of science.


2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yuu Asano ◽  
Kensuke Yamashita ◽  
Aoi Hasegawa ◽  
Takanori Ogasawara ◽  
Hoshie Iriki ◽  
...  

AbstractThe powerful genome editing tool Streptococcus pyogenes Cas9 (SpCas9) requires the trinucleotide NGG as a protospacer adjacent motif (PAM). The PAM requirement is limitation for precise genome editing such as single amino-acid substitutions and knock-ins at specific genomic loci since it occurs in narrow editing window. Recently, SpCas9 variants (i.e., xCas9 3.7, SpCas9-NG, and SpRY) were developed that recognise the NG dinucleotide or almost any other PAM sequences in human cell lines. In this study, we evaluated these variants in Dictyostelium discoideum. In the context of targeted mutagenesis at an NG PAM site, we found that SpCas9-NG and SpRY were more efficient than xCas9 3.7. In the context of NA, NT, NG, and NC PAM sites, the editing efficiency of SpRY was approximately 60% at NR (R = A and G) but less than 22% at NY (Y = T and C). We successfully used SpRY to generate knock-ins at specific gene loci using donor DNA flanked by 60 bp homology arms. In addition, we achieved point mutations with efficiencies as high as 97.7%. This work provides tools that will significantly expand the gene loci that can be targeted for knock-out, knock-in, and precise point mutation in D. discoideum.


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