Targeted mutagenesis in rice using CRISPR-Cas system

Miao, Junwei; Guo, Dongshu; Zhang, Jinzhe; Qing, Huang; Qin, Genji; Zhang, Xin; Wan, Jianmin; Gu, Hongya; Liu, Qu

doi:10.1038/cr.2013.123

Cited by 770 publications

(524 citation statements)

References 13 publications

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“…Since 2013, CRISPR/Cas9 has been successfully applied by transient expression and/or stable transgenic lines in several plant species, such as Arabidopsis, Nicotiana benthamiana, rice, wheat, maize, and tomato (Brooks et al 2014;Jiang et al 2013;Li et al 2013;Miao et al 2013;Nekrasov et al 2013;Shan et al 2013). Moreover, the mutations generated in the primary transgenic plants by the CRISPR/Cas9 system can be stably transmitted to the next generation (Brooks et al 2014;Feng et al 2014).…”

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confidence: 99%

Efficient targeted mutagenesis in potato by the CRISPR/Cas9 system

et al. 2015

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confidence: 99%

Efficient targeted mutagenesis in potato by the CRISPR/Cas9 system

et al. 2015

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“…It was shown that engineered CRISPR/Cas caused mutations in target genes or corrections in transgenes in transient expression assays in plant protoplasts and tobacco leaves (10). Importantly, stable expression of the CRISPR/ Cas in transgenic Arabidopsis, tobacco, and rice plants led to mutations (mostly indels) in target genes and correction of a transgene (4)(5)(6)(7)(8)(9). However, it was not known whether the gene mutations and corrections occurred in somatic cells only or whether some of the mutations and corrections happened in germ-line cells and thus may be heritable.…”

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confidence: 99%

“…The CRISPR (clustered regularly interspaced short palindromic repeat)/Cas (CRISPR-associated) system has been successfully used for efficient genome editing in human cell lines, zebrafish, and mouse (1)(2)(3) and recently applied to gene modification in plants (4)(5)(6)(7)(8)(9)(10). In this system a short RNA molecule guides the associated endonuclease Cas9 to generate double strand breaks (DSBs) in the target genomic DNA, which lead to sequence mutations as a result of error-prone nonhomologous end-joining (NHEJ) DNA damage repair or to gene correction or replacement as a result of homologydependent recombination (HR) (11).…”

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confidence: 99%

Multigeneration analysis reveals the inheritance, specificity, and patterns of CRISPR/Cas-induced gene modifications in Arabidopsis

Feng

Mao

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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The CRISPR (clustered regularly interspaced short palindromic repeat)/Cas (CRISPR-associated) system has emerged as a powerful tool for targeted gene editing in many organisms, including plants. However, all of the reported studies in plants focused on either transient systems or the first generation after the CRISPR/ Cas system was stably transformed into plants. In this study we examined several plant generations with seven genes at 12 different target sites to determine the patterns, efficiency, specificity, and heritability of CRISPR/Cas-induced gene mutations or corrections in Arabidopsis. The proportion of plants bearing any mutations (chimeric, heterozygous, biallelic, or homozygous) was 71.2% at T1, 58.3% at T2, and 79.4% at T3 generations. CRISPR/Cas-induced mutations were predominantly 1 bp insertion and short deletions. Gene modifications detected in T1 plants occurred mostly in somatic cells, and consequently there were no T1 plants that were homozygous for a gene modification event. In contrast, ∼22% of T2 plants were found to be homozygous for a modified gene. All homozygotes were stable to the next generation, without any new modifications at the target sites. There was no indication of any off-target mutations by examining the target sites and sequences highly homologous to the target sites and by in-depth whole-genome sequencing. Together our results show that the CRISPR/Cas system is a useful tool for generating versatile and heritable modifications specifically at target genes in plants.

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“…using CRISPR/Cas-based genome editing in plants and in animals (3)(4)(5)(6) To use CRISPR/Cas for crop improvement, mutations generated by CRISPR/Cas have to be stable and heritable. Previous CRISPR studies in plant systems focused on testing the feasibility of the CRISPR/Cas system and were performed only on cultured cells or on the first generation of the transgenic lines (4,5,7).…”

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confidence: 99%