Efficient CRISPR/Cas9‐based plant genomic fragment deletions by microhomology‐mediated end joining

Tan, Jiantao; Zhao, Yongchun; Wang, Bin; Yu, Hao; Wang, Yaxi; Li, Yangyang; Luo, Wenjun; Zong, Wubei; Li, Gousi; Chen, Shuifu; Ma, Kun; Xie, Xianrong; Chen, Letian; Liu, Yao-Guang; Zhu, Qinlong

doi:10.1111/pbi.13390

Cited by 39 publications

(39 citation statements)

References 10 publications

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“…Very recently, MMEJ was also shown to be dominant among DSB repaired outcomes of the CRISPR/Cas‐based genome editing in plants (Tan et al ., 2020; Weiss et al ., 2020) and could be engineered for precise deletions of plant genomic DNAs (Tan et al ., 2020) or even chromosomal translocations (Beying et al ., 2020). MMEJ efficiently involved in repairing DSBs generated by CRISPR/Cas complexes resulted in DNA fragment deletion ranging from some dozens of base pairs to more than 20 kb (Tan et al ., 2020) or heritable chromosomal translocations in the Mbp range in Arabidopsis (Beying et al ., 2020). The frequencies of CRISPR/Cas‐based DSB repair by MMEJ were similar to that of cNHEJ and more efficient in the absence of the cNHEJ pathway (Beying et al ., 2020; Tan et al ., 2020; Weiss et al ., 2020).…”

Section: Engineering Mmej For Precision Gene Editingmentioning

confidence: 99%

“…Harnessing the favourable conditions for MMEJ would further improve its precision editing. It would be useful for plant engineers to determine and optimize the conditions for MMEJ‐based editing since little information has been released about its applications in plants (Beying et al ., 2020; Tan et al ., 2020; Weiss et al ., 2020).…”

Section: Engineering Mmej For Precision Gene Editingmentioning

confidence: 99%

“…MMEJ‐based gene editing offers alternative tools for precision engineering of organisms of interest at a potentially higher frequency than what is achieved with HR‐based approaches. Substantial understanding of MMEJ activation and its mechanism in DSB repair (Figure 1) has been made in animal and plant studies (Ata et al ., 2018; Bae et al ., 2014; Beying et al ., 2020; Sfeir and Symington, 2015; Tan et al ., 2020; Weiss et al ., 2020). Therefore, subsequent applications in precision gene knock‐in using animals showed significantly higher efficiencies than that of HR in the same experimental conditions.…”

Section: Concluding Remarks and Future Perspectivesmentioning

confidence: 99%

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CRISPR/Cas‐based precision genome editing via microhomology‐mediated end joining

Doan

Kim

et al. 2020

Plant Biotechnology Journal

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Summary Gene editing and/or allele introgression with absolute precision and control appear to be the ultimate goals of genetic engineering. Precision genome editing in plants has been developed through various approaches, including oligonucleotide‐directed mutagenesis (ODM), base editing, prime editing and especially homologous recombination (HR)‐based gene targeting. With the advent of CRISPR/Cas for the targeted generation of DNA breaks (single‐stranded breaks (SSBs) or double‐stranded breaks (DSBs)), a substantial advancement in HR‐mediated precise editing frequencies has been achieved. Nonetheless, further research needs to be performed for commercially viable applications of precise genome editing; hence, an alternative innovative method for genome editing may be required. Within this scope, we summarize recent progress regarding precision genome editing mediated by microhomology‐mediated end joining (MMEJ) and discuss their potential applications in crop improvement.

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Section: Engineering Mmej For Precision Gene Editingmentioning

confidence: 99%

Section: Engineering Mmej For Precision Gene Editingmentioning

confidence: 99%

Section: Concluding Remarks and Future Perspectivesmentioning

confidence: 99%

See 1 more Smart Citation

CRISPR/Cas‐based precision genome editing via microhomology‐mediated end joining

Doan

Kim

et al. 2020

Plant Biotechnology Journal

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“…NHEJ is the preferred repair pathway, because it does not require a homologous repair template [22]. If the target sequence is near microhomologous sequences, then the DSBs may be repaired through microhomology-mediated end-joining, resulting in fragment deletions of the plant genome with higher efficiency [23,24].…”

Section: Introductionmentioning

confidence: 99%

The Development of Herbicide Resistance Crop Plants Using CRISPR/Cas9-Mediated Gene Editing

Dong

Huang

Wang

2021

Genes

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The rapid increase in herbicide-resistant weeds creates a huge challenge to global food security because it can reduce crop production, causing considerable losses. Combined with a lack of novel herbicides, cultivating herbicide-resistant crops becomes an effective strategy to control weeds because of reduced crop phytotoxicity, and it expands the herbicidal spectrum. Recently developed clustered regularly interspaced short palindromic repeat/CRISPR-associated protein (CRISPR/Cas)-mediated genome editing techniques enable efficiently targeted modification and hold great potential in creating desired plants with herbicide resistance. In the present review, we briefly summarize the mechanism responsible for herbicide resistance in plants and then discuss the applications of traditional mutagenesis and transgenic breeding in cultivating herbicide-resistant crops. We mainly emphasize the development and use of CRISPR/Cas technology in herbicide-resistant crop improvement. Finally, we discuss the future applications of the CRISPR/Cas system for developing herbicide-resistant crops.

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“…For example, SpCas9 nuclease can be used to genetically modify a specific target locus within the plant genome. Briefly, SpCas9 catalyzes a double-stranded break (DSB) which subsequently stimulates diverse DNA repair mechanisms [non-homologous end joining (NHEJ), microhomology-mediated end joining (MMEJ), and homology-directed repairs (HDR)] resulting in gene knockout, DNA fragment insertion, deletion, and replacement as specifically required [2][3][4][5][6]. Alternatively, the SpCas9(D10A) nickase can be utilized to drive various engineered nucleoside deaminases to the target region and catalyze the hydrolytic deamination of cytosines to uracils and/or adenosines to inosine within the editing window.…”

Section: Introductionmentioning

confidence: 99%

SpRY greatly expands the genome editing scope in rice with highly flexible PAM recognition

Kuang

Ren

et al. 2020

Preprint

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Plant genome engineering mediated by various CRISPR-based tools requires specific protospacer adjacent motifs (PAMs), such as the well-performed NGG, NG, NNG, etc., to initiate the target recognition, which notably restricts the editable range of the plant genome. In this study, we thoroughly investigated the nuclease activity and the PAM preference of two structurally-engineered SpCas9 variants (SpG and SpRY) in transgenic rice. Our study shows that SpG nuclease favors NGD PAMs, albeit less efficiently than the previously described SpCas9-NG and that SpRY nuclease achieves efficient editing across a wide range of genomic loci, exhibiting a preference of NGD as well as NAN PAMs. Furthermore, SpRY-fused cytidine deaminase hAID*Δ and adenosine deaminase TadA8e were generated, respectively. These constructs efficiently induced C-to-T and A-to-G conversions in the target genes toward various non-canonical PAMs, including non-G PAMs. Remarkably, high-frequency self-editing events (indels and DNA fragments deletion) in the integrated T-DNA fragments as a result of the nuclease activity of SpRY were observed, whereas the self-editing of SpRY nickase-mediated base editor was quite low in transgenic rice lines. In conclusion, the broad PAM compatibility of SpRY greatly expands the targeting scope of CRISPR-based tools in plant genome engineering.

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Efficient CRISPR/Cas9‐based plant genomic fragment deletions by microhomology‐mediated end joining

Cited by 39 publications

References 10 publications

CRISPR/Cas‐based precision genome editing via microhomology‐mediated end joining

CRISPR/Cas‐based precision genome editing via microhomology‐mediated end joining

The Development of Herbicide Resistance Crop Plants Using CRISPR/Cas9-Mediated Gene Editing

SpRY greatly expands the genome editing scope in rice with highly flexible PAM recognition

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