Discovery of rice essential genes by characterizing a CRISPR‐edited mutation of closely related rice MAP kinase genes

Minkenberg, Bastian; Xie, Kabin; Yang, Yinong

doi:10.1111/tpj.13399

Cited by 68 publications

(48 citation statements)

References 57 publications

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“…Multiplex PTG/Cas9 systems can help with multigene family analysis, as reported for the closely related mitogen-activated protein kinase MPK1 and MPK6 in rice (Minkenberg et al, 2017). 67% of all lines were double mutants for MPK genes with a high frequency of biallelic mutations on multiple target sites.…”

Section: Multiplex Genome Editing: When Does It Become Useful?mentioning

confidence: 99%

The Enhancement of Plant Disease Resistance Using CRISPR/Cas9 Technology

et al. 2018

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Genome editing technologies have progressed rapidly and become one of the most important genetic tools in the implementation of pathogen resistance in plants. Recent years have witnessed the emergence of site directed modification methods using meganucleases, zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and clustered regularly interspaced short palindrome repeats (CRISPR)/CRISPR-associated protein 9 (Cas9). Recently, CRISPR/Cas9 has largely overtaken the other genome editing technologies due to the fact that it is easier to design and implement, has a higher success rate, and is more versatile and less expensive. This review focuses on the recent advances in plant protection using CRISPR/Cas9 technology in model plants and crops in response to viral, fungal and bacterial diseases. As regards the achievement of viral disease resistance, the main strategies employed in model species such as Arabidopsis and Nicotiana benthamiana, which include the integration of CRISPR-encoding sequences that target and interfere with the viral genome and the induction of a CRISPR-mediated targeted mutation in the host plant genome, will be discussed. Furthermore, as regards fungal and bacterial disease resistance, the strategies based on CRISPR/Cas9 targeted modification of susceptibility genes in crop species such as rice, tomato, wheat, and citrus will be reviewed. After spending years deciphering and reading genomes, researchers are now editing and rewriting them to develop crop plants resistant to specific pests and pathogens.

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Section: Multiplex Genome Editing: When Does It Become Useful?mentioning

confidence: 99%

The Enhancement of Plant Disease Resistance Using CRISPR/Cas9 Technology

et al. 2018

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“…The start and end sites of the tRNA in the tandemly arrayed tRNA‐sgRNA transcripts are precisely recognized and cleaved by endogenous RNases (RNase P and RNase Z in plants) to simultaneously produce multiple functional sgRNAs (Xie et al ., ). Additionally, the tRNA gene contains internal promoter elements, suggesting that it may work as a potential transcriptional enhancer, as verified in a recent study involving rice, which showed that the relative expression level of the sgRNA in the PTG/Cas9 system is higher than in the CRISPR/Cas9 system (Minkenberg et al ., ; Xie et al ., ). Compared to the traditional CRISPR/Cas9 system, the PTG/Cas9 system is simpler and more efficient for multiplex targeted genome modifications.…”

Section: Introductionmentioning

confidence: 99%

Optimized paired‐sgRNA/Cas9 cloning and expression cassette triggers high‐efficiency multiplex genome editing in kiwifruit

Wang

et al. 2018

Plant Biotechnology Journal

138

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SummaryKiwifruit is an important fruit crop; however, technologies for its functional genomic and molecular improvement are limited. The clustered regulatory interspaced short palindromic repeats (CRISPR)/CRISPR‐associated protein (Cas) system has been successfully applied to genetic improvement in many crops, but its editing capability is variable depending on the different combinations of the synthetic guide RNA (sgRNA) and Cas9 protein expression devices. Optimizing conditions for its use within a particular species is therefore needed to achieve highly efficient genome editing. In this study, we developed a new cloning strategy for generating paired‐sgRNA/Cas9 vectors containing four sgRNAs targeting the kiwifruit phytoene desaturase gene (AcPDS). Comparing to the previous method of paired‐sgRNA cloning, our strategy only requires the synthesis of two gRNA‐containing primers which largely reduces the cost. We further compared efficiencies of paired‐sgRNA/Cas9 vectors containing different sgRNA expression devices, including both the polycistronic tRNA‐sgRNA cassette (PTG) and the traditional CRISPR expression cassette. We found the mutagenesis frequency of the PTG/Cas9 system was 10‐fold higher than that of the CRISPR/Cas9 system, coinciding with the relative expressions of sgRNAs in two different expression cassettes. In particular, we identified large chromosomal fragment deletions induced by the paired‐sgRNAs of the PTG/Cas9 system. Finally, as expected, we found both systems can successfully induce the albino phenotype of kiwifruit plantlets regenerated from the G418‐resistance callus lines. We conclude that the PTG/Cas9 system is a more powerful system than the traditional CRISPR/Cas9 system for kiwifruit genome editing, which provides valuable clues for optimizing CRISPR/Cas9 editing system in other plants.

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“…Therefore, the use of proxies of gene editing in the T2 progenies could 117 significantly reduce the time and effort invested in identifying CRISPR-mutagenized 118 plants. 119 CRISPR/Cas9 can simultaneously edit multiple loci (co-editing) at high frequency in the 120 somatic tissues of T1 Arabidopsis and rice plants (Ma et al 2015;Yan et al 2016;121 Zhang et al 2016;Minkenberg et al 2017). Hence, we hypothesized that we could take 122 advantage of this high co-editing frequency to aid in the selection of CRISPR-123 mutagenized plants.…”

Section: Introduction 46mentioning

confidence: 99%

Proxies of CRISPR/Cas9 Activity To Aid in the Identification of Mutagenized Arabidopsis Plants

Vavrik

Danna

2020

G3 Genes|Genomes|Genetics

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24CRISPR/Cas9 has become the preferred gene-editing technology to obtain loss-of-25 function mutants in plants, and hence a valuable tool to study gene function. This is 26 mainly due to the easy reprogramming of Cas9 specificity using customizable small 27 non-coding RNAs, and to the possibility of editing several independent genes 28 simultaneously. Despite these advances, the identification of CRISPR-edited plants 29remains time and resource-intensive. Here, based on the premise that one editing event 30in one locus is a good predictor of editing event/s in other locus/loci, we developed a 31 CRISPR co-editing selection strategy that greatly facilitates the identification of 32 CRISPR-mutagenized Arabidopsis thaliana plants. This strategy is based on targeting 33 the gene/s of interest simultaneously with a proxy of CRISPR-Cas9-directed 34 mutagenesis. The proxy is an endogenous gene whose loss-of-function produces an 35 easy-to-detect visible phenotype that is unrelated to the expected phenotype of the 36 gene/s under study. We tested this strategy via assessing the frequency of co-editing of 37 three functionally unrelated proxy genes. We found that each proxy predicted the 38 occurrence of mutations in each surrogate gene with efficiencies ranging from 68% to 39

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Discovery of rice essential genes by characterizing a CRISPR‐edited mutation of closely related rice MAP kinase genes

Cited by 68 publications

References 57 publications

The Enhancement of Plant Disease Resistance Using CRISPR/Cas9 Technology

The Enhancement of Plant Disease Resistance Using CRISPR/Cas9 Technology

Optimized paired‐sgRNA/Cas9 cloning and expression cassette triggers high‐efficiency multiplex genome editing in kiwifruit

Proxies of CRISPR/Cas9 Activity To Aid in the Identification of Mutagenized Arabidopsis Plants

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