Demonstration of CRISPR/Cas9/sgRNA-mediated targeted gene modification in Arabidopsis, tobacco, sorghum and rice

By W. Jiang, Huanbin Zhou, Honghao Bi, M. Fromm, Bing Yang, D. Weeks. Nucleic Acids Research, 2013

1,148 citations recorded by the source corpus.

The type II CRISPR/Cas system from Streptococcus pyogenes and its simplified derivative, the Cas9/single guide RNA (sgRNA) system, have emerged as potent new tools for targeted gene knockout in bacteria, yeast, fruit fly, zebrafish and human cells. Here, we describe adaptations of these systems leading to successful expression of the Cas9/sgRNA system in two dicot plant species, Arabidopsis and tobacco, and two monocot crop species, rice and sorghum. Agrobacterium tumefaciens was used for delivery of genes encoding Cas9, sgRNA and a non-fuctional, mutant green fluorescence protein (GFP) to Arabidopsis and tobacco. The mutant GFP gene contained target sites in its 5′ coding regions that were successfully cleaved by a CAS9/sgRNA complex that, along with error-prone DNA repair, resulted in creation of functional GFP genes. DNA sequencing confirmed Cas9/sgRNA-mediated mutagenesis at the target site. Rice protoplast cells transformed with Cas9/sgRNA constructs targeting the promoter region of the bacterial blight susceptibility genes, OsSWEET14 and OsSWEET11, were confirmed by DNA sequencing to contain mutated DNA sequences at the target sites. Successful demonstration of the Cas9/sgRNA system in model plant and crop species bodes well for its near-term use as a facile and powerful means of plant genetic engineering for scientific and agricultural applications.

Subfields: Molecular Biology, Plant Science

View Paper: https://www.semanticscholar.org/paper/92d7f5432b123198b7f88f4f9b1bd83bce493f76

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Citations in the corpus

The CRISPR/Cas9 system for plant genome editing and beyond. (2015, background): “Subsequent work focused on additional crop species such as sorghum (Jiang et al., 2013b), wheat (Upadhyay et al., 2013; Wang et al., 2014b) andmaize (Liang et al., 2014).”
A CRISPR/Cas9 toolkit for multiplex genome editing in plants (2014, background): “The CRISPR/Cas system has been harnessed to achieve efficient genome editing in a variety of organisms, including bacteria, yeast, plants, and animals, as well as human cell lines [12-27].”
The Sequence of 1504 Mutants in the Model Rice Variety Kitaake Facilitates Rapid Functional Genomic Studies (2017, background): “, 2012), CRISPR/Cas9 genome editing (Jiang et al., 2013; Miao et al., 2013; Xie et al., 2015), chemical induction, such as ethyl methanesulfonate (EMS) (Henry et al.”
A Robust CRISPR/Cas9 System for Convenient, High-Efficiency Multiplex Genome Editing in Monocot and Dicot Plants. (2015, methodology): “For Gibson Assembly cloning, home-made 23 isothermal in vitro recombination master mixture was prepared as previously described (Jiang et al., 2013a, 2013b).”
sgRNAcas9: A Software Package for Designing CRISPR sgRNA and Evaluating Potential Off-Target Cleavage Sites (2014, neutral): “upstream of the PAM site [34].”
CRISPR/Cas9 in Genome Editing and Beyond. (2016, background): “The use of the Cas9 platform has greatly increased the efficiency of generating transgenic organisms, from fungi (32) and plants (33, 34, 97, 98) to a variety of animals (5–7, 36, 99–102) (reviewed in 9, 10, 35).”
Enhanced Rice Blast Resistance by CRISPR/Cas9-Targeted Mutagenesis of the ERF Transcription Factor Gene OsERF922 (2016, background): “In particular, CRISPR/Cas9 has been demonstrated to be the most effective SSN thus far and has been used for genome editing in major crops such as rice [29–41], maize [42–45], wheat [39, 46, 47], sorghum [30], tomato [48, 49], soybean [50–52] and potato [53].”
Efficient Gene Editing in Tomato in the First Generation Using the Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR-Associated9 System1 (2014, background): “…Arabidopsis (Arabidopsis thaliana; Li et al., 2013) and Nicotiana benthamiana (Nekrasov et al., 2013), there have also been reports for three crop species, rice (Oryza sativa; Zhang et al., 2014), sorghum (Sorghum bicolor; Jiang et al., 2013), and wheat (Triticum aestivum; Wang et al., 2014).”
Egg cell-specific promoter-controlled CRISPR/Cas9 efficiently generates homozygous mutants for multiple target genes in Arabidopsis in a single generation (2015, neutral): “system provides an alternative method [17–44].”
Gene Pyramiding for Sustainable Crop Improvement against Biotic and Abiotic Stresses (2020, background): “Recurrent selection is considered an efficient approach for pyramiding multiple traits in plants; however, its efficacy of selection is not satisfactory because phenotypic selection depends on environments while genotypic selection takes much time (two to three cropping seasons for a cycle of select”