Large chromosomal deletions and heritable small genetic changes induced by CRISPR/Cas9 in rice

Zhou, Huanbin; Liu, Bo; Weeks, Donald P.; Spalding, Martin H.; Yang, Bing

doi:10.1093/nar/gku806

Cited by 536 publications

(454 citation statements)

References 47 publications

(62 reference statements)

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“…Highly efficient, stable mutagenesis was subsequently demonstrated in Arabidopsis, as well as in several crops: rice, wheat, barley, Brassica oleracea , potato, maize and soybean (Brooks et al ., 2014; Butler et al ., 2015; Čermák et al ., 2015; Fauser et al ., 2014; Feng et al ., 2014; Ito et al ., 2015; Jiang et al ., 2014; Lawrenson et al ., 2015; Li et al ., 2015; Svitashev et al ., 2015; Wang et al ., 2014; Zhang et al ., 2014; Zhou et al ., 2014). The CRISPR‐Cas9 system can be used to achieve multiple mutagenesis by the expression of several sgRNAs targetting different genes (Lowder et al ., 2015; Ma et al ., 2015).…”

Section: Introductionmentioning

confidence: 99%

Selective gene dosage by CRISPR‐Cas9 genome editing in hexaploid Camelina sativa

Morineau

Bellec

Tellier

et al. 2017

Plant Biotechnology Journal

244

166

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In many plant species, gene dosage is an important cause of phenotype variation. Engineering gene dosage, particularly in polyploid genomes, would provide an efficient tool for plant breeding. The hexaploid oilseed crop Camelina sativa, which has three closely related expressed subgenomes, is an ideal species for investigation of the possibility of creating a large collection of combinatorial mutants. Selective, targeted mutagenesis of the three delta‐12‐desaturase (FAD2) genes was achieved by CRISPR‐Cas9 gene editing, leading to reduced levels of polyunsaturated fatty acids and increased accumulation of oleic acid in the oil. Analysis of mutations over four generations demonstrated the presence of a large variety of heritable mutations in the three isologous CsFAD2 genes. The different combinations of single, double and triple mutants in the T3 generation were isolated, and the complete loss‐of‐function mutants revealed the importance of delta‐12‐desaturation for Camelina development. Combinatorial association of different alleles for the three FAD2 loci provided a large diversity of Camelina lines with various lipid profiles, ranging from 10% to 62% oleic acid accumulation in the oil. The different allelic combinations allowed an unbiased analysis of gene dosage and function in this hexaploid species, but also provided a unique source of genetic variability for plant breeding.

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Section: Introductionmentioning

confidence: 99%

Selective gene dosage by CRISPR‐Cas9 genome editing in hexaploid Camelina sativa

Morineau

Bellec

Tellier

et al. 2017

Plant Biotechnology Journal

244

166

View full text Add to dashboard Cite

show abstract

“…This improvement further simplifies the CRISPR/Cas9 system and enhances reagent delivery. After initial reports, many studies quickly announced successful and effective CRISPR/Cas9-mediated genome editing in plants (Feng et al, 2013Jiang et al, 2013;Li et al, 2013;Mao et al, 2013;Miao et al, 2013;Nekrasov et al, 2013;Shan et al, 2013b;Xie and Yang, 2013;Fauser et al, 2014;Schiml et al, 2014;Zhou et al, 2014). Due to simplified engineering of target specificity and its dual-component nature, CRISPR/ Cas9 allows for simultaneous targeting of multiple genomic loci.…”

mentioning

confidence: 99%

A CRISPR/Cas9 Toolbox for Multiplexed Plant Genome Editing and Transcriptional Regulation

et al. 2015

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The relative ease, speed, and biological scope of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPRassociated Protein9 (Cas9)-based reagents for genomic manipulations are revolutionizing virtually all areas of molecular biosciences, including functional genomics, genetics, applied biomedical research, and agricultural biotechnology. In plant systems, however, a number of hurdles currently exist that limit this technology from reaching its full potential. For example, significant plant molecular biology expertise and effort is still required to generate functional expression constructs that allow simultaneous editing, and especially transcriptional regulation, of multiple different genomic loci or multiplexing, which is a significant advantage of CRISPR/Cas9 versus other genome-editing systems. To streamline and facilitate rapid and wide-scale use of CRISPR/Cas9-based technologies for plant research, we developed and implemented a comprehensive molecular toolbox for multifaceted CRISPR/Cas9 applications in plants. This toolbox provides researchers with a protocol and reagents to quickly and efficiently assemble functional CRISPR/Cas9 transfer DNA constructs for monocots and dicots using Golden Gate and Gateway cloning methods. It comes with a full suite of capabilities, including multiplexed gene editing and transcriptional activation or repression of plant endogenous genes. We report the functionality and effectiveness of this toolbox in model plants such as tobacco (Nicotiana benthamiana), Arabidopsis (Arabidopsis thaliana), and rice (Oryza sativa), demonstrating its utility for basic and applied plant research.

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“…The components of the prokaryotic clustered, regularly interspaced, short palindromic repeats (CRISPR)/ CRISPR-associated (Cas) system is a recently developed technology for targeted genome modification in mammalian cells, bacteria, zebra fish, mice, monkey and pigs [15][16][17][18]. When these indels introduce a frame shift mutation or disrupt important functional domains, the functions of the target genes will be damaged [19][20][21]. Co-injection of zygotes with Cas9 mRNA and single guide (sg) RNA has proven to be an efficient gene-editing strategy.…”

Section: Introductionmentioning

confidence: 99%

Double-Muscled Phenotype in Mutant Sheep Directed by the CRISPRCas9 System

Wu¹,

Du²,

Liu³

et al. 2018

Clon Transgen

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Myostatin (MSTN) is a well-known negative regulator of muscle growth. The double-muscled sheep caused by natural loss-of-function mutations of MSTN have very strong skeletal muscle. In this study, our results demonstrate the successful generation of MSTN mutant sheep via specific targeting of an exon 1 site using Cas9 technology. The MSTN-knockout sheep in our study had increased muscle significantly just like double-muscled phenotype. Our study suggests that the direct injection of Cas9: sgRNA into zygotes could be widely used to create gene knockouts in large domestic animals. Notably, on the basis of our findings, sheep can be added to the growing list of species for which genome editing is now practical. The generation of MSTN mutant sheep has implications for the genetic improvement of local sheep varieties, and also for the usage of sheep as a model for large animal medical research.

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Large chromosomal deletions and heritable small genetic changes induced by CRISPR/Cas9 in rice

Cited by 536 publications

References 47 publications

Selective gene dosage by CRISPR‐Cas9 genome editing in hexaploid Camelina sativa

Selective gene dosage by CRISPR‐Cas9 genome editing in hexaploid Camelina sativa

A CRISPR/Cas9 Toolbox for Multiplexed Plant Genome Editing and Transcriptional Regulation

Double-Muscled Phenotype in Mutant Sheep Directed by the CRISPRCas9 System

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