Activity and specificity of TRV-mediated gene editing in plants

Ali, Zahir; Abulfaraj, Aala A.; Piatek, Marek J.; Mahfouz, Magdy M.

doi:10.1080/15592324.2015.1044191

Cited by 65 publications

(48 citation statements)

References 21 publications

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“…Further, data indicate that TRV-mediated genome editing exhibited no off-target activities at potential off-targets indicating the precision of the system for plant genome engineering. Taken together, the data establish the feasibility and exciting possibilities of using virus-mediated CRISPR/Cas9 for targeted engineering of plant genomes (Ali et al, 2015).…”

Section: Delivery Of Constructs Into Target Organismmentioning

confidence: 63%

Genome editing for crop improvement: Challenges and opportunities

2015

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ABSTRACT. Genome or gene editing includes several new techniques to help scientists precisely modify genome sequences. The techniques also enables us to alter the regulation of gene expression patterns in a pre-determined region and facilitates novel insights into the functional genomics of an organism.Emergence of genome editing has brought considerable excitement especially among agricultural scientists because of its simplicity, precision and power as it offers new opportunities to develop improved crop varieties with clear-cut addition of valuable traits or removal of undesirable traits. Research is underway to improve crop varieties with higher yields, strengthen stress tolerance, disease and pest resistance, decrease input costs, and increase nutritional value. Genome editing encompasses a wide variety of tools using either a site-specific recombinase (SSR) or a site-specific nuclease (SSN) system. Both systems require recognition of a known sequence. The SSN system generates single or double strand DNA breaks and activates endogenous DNA repair pathways. SSR technology, such as Cre/loxP and Flp/FRT mediated systems, are able to knockdown or knock-in genes in the genome of eukaryotes, depending on the orientation of the specific sites (loxP, FLP, etc.) flanking the target site. There are 4 main classes of SSN developed to cleave genomic sequences, mega-nucleases (homing endonuclease), zinc finger nucleases (ZFNs), transcriptional activator-like effector nucleases (TALENs), and the CRISPR/Cas nuclease system (clustered regularly interspaced short palindromic repeat/CRISPR-associated protein). The recombinase mediated genome engineering depends on recombinase (sub-) family and target-site and induces high frequencies of homologous recombination.

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Section: Delivery Of Constructs Into Target Organismmentioning

confidence: 63%

Genome editing for crop improvement: Challenges and opportunities

2015

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“…To avoid the necessity for multigene constructs and to increase the expression level of the sgRNA, viral delivery vectors have been used to transiently express sgRNAs in plants overexpressing Cas9 from a stably integrated transgene. Both Tobacco Rattle Virus (TRV) [29] and Cabbage Leaf Curl Virus (CaLCuV) [30] have been adapted for this purpose. More recently, Cody et al [31] demonstrated targeted mutagenesis induced by transient expression of Cas9 simultaneous with delivery of sgRNAs from a tobacco mosaic virus-derived vector.…”

Section: Toolkits For Targeted Mutagenesismentioning

confidence: 99%

CRISPR-based tools for plant genome engineering

Silva

Patron

2017

Emerging Topics in Life Sciences

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Molecular tools adapted from bacterial CRISPR (clustered regulatory interspaced short palindromic repeat) adaptive immune systems have been demonstrated in an increasingly wide range of plant species. They have been applied for the induction of targeted mutations in one or more genes as well as for directing the integration of new DNA to specific genomic loci. The construction of molecular tools for multiplexed CRISPR-mediated editing in plants has been facilitated by cloning techniques that allow multiple sequences to be assembled together in a single cloning reaction. Modifications of the canonical Cas9 protein from Streptococcus pyogenes and the use of nucleases from other bacteria have increased the diversity of genomic sequences that can be targeted and allow the delivery of protein cargos such as transcriptional activators and repressors. Furthermore, the direct delivery of protein-RNA complexes to plant cells and tissues has enabled the production of engineered plants without the delivery or genomic integration of foreign DNA. Here, we review toolkits derived from bacterial CRISPR systems for targeted mutagenesis, gene delivery and modulation of gene expression in plants, focusing on their composition and the strategies employed to reprogramme them for the recognition of specific genomic targets.

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“…Oligonucleotide-mediated technology to target different genes in Arabidopsis thaliana (hereafter, Arabidopsis) protoplasts showed that the selection of traits in plant varieties may improve crop performance by genome editing technologies (Sauer et al 2015). A virus-based guide RNA (gRNA) delivery system for CRISPR/Cas9-mediated plant genome editing (VIGE) has been used in Nicotiana benthamiana, fruit trees, and crop plants (Ali et al 2015;Ilardi and Tavazza 2015;Yin et al 2015). Using a modified cabbage leaf curl virus (CaLCuV) vector to express gRNAs in stable transgenic plants expressing Cas9, mutations in NbPDS3 and NbIspH genes were confirmed by DNA sequencing, indicating that geminivirus-based VIGE could be a powerful tool in plant genome editing (Yin et al 2015).…”

Section: Delivery Of the Crispr/cas9 System In Plantsmentioning

confidence: 99%

“…Plum pox virus (PPV) is the etiological agent of sharka, the most devastating and economically important viral disease affecting Prunus species. Robust and predictable resistance can be achieved using CRISPR/Cas9-engineered nucleases to target conserved regions of the PPV genome and knockout host susceptibility genes in stone fruit trees for efficient and fast breeding programs (Ali et al 2015;Cermak et al 2015;Ilardi and Tavazza 2015;Yin et al 2015). The results of tobacco rattle virus (TRV)-mediated genome editing in Nicotiana benthamiana demonstrated the persistence of the TRV-mediated Cas9 activity for up to 30 d after agroinfection, indicating that virus-mediated CRISPR/Cas9 is a useful tool for targeted engineering of plant genomes (Ali et al 2015).…”

Section: Delivery Of the Crispr/cas9 System In Plantsmentioning

confidence: 99%

“…To streamline and facilitate rapid and wide-scale use of CRISPR/Cas9-based technologies for plant research, a comprehensive molecular toolbox for multifaceted CRISPR/Cas9 applications was developed for Arabidopsis for multiplexed gene editing and transcriptional activation or repression of plant endogenous genes (Ali et al 2015;Lowder et al 2015). With this system, uniform biallelic, heterozygous, homozygous, and chimeric mutations in Arabidopsis T1 plants have been obtained, and the targeted mutations in Arabidopsis were heritable; thus, a versatile toolbox is available for studying functions of multiple genes and gene families in plants for basic research and genetic improvement Sauer et al 2015).…”

Section: Arabidopsis Thalianamentioning

confidence: 99%

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Applications and roles of the CRISPR system in genome editing of plants

Tang

2016

J. For. Res.

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Genome editing is a valuable tool to target specific DNA sequences for mutagenesis in the genomes of microbes, plants, and animals. Although different genome editing technologies are available, the clustered regularly interspaced short palindromic repeats/Cas9 (CRISPR/ Cas9) system, which utilizes engineered endonucleases to generate a double-stranded DNA break (DSB) in the target DNA region and subsequently stimulates site-specific mutagenesis through DNA repair machineries, is emerging as a powerful genome editing tool for elucidating mechanisms of protection from plant viruses, plant disease resistance, and gene functions in basic and applied research. In this review, we provide an overview of recent advances in the CRISPR system associated genome editing in plants by focusing on application of this technology in model plants, crop plants, fruit plants, woody plants and grasses and discuss how genome editing associated with the CRISPR system can provide insights into genome modifications and functional genomics in plants.

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Activity and specificity of TRV-mediated gene editing in plants

Cited by 65 publications

References 21 publications

Genome editing for crop improvement: Challenges and opportunities

Genome editing for crop improvement: Challenges and opportunities

CRISPR-based tools for plant genome engineering

Applications and roles of the CRISPR system in genome editing of plants

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