High-efficiency prime editing with optimized, paired pegRNAs in plants

Lin, Qiupeng; Jin, Shuai; Zong, Yuan; Yu, Hong; Zhu, Zixu; Liu, Guanwen; Kou, Liquan; Wang, Yanpeng; Qiu, Jian; Li, Jiayang; Gao, Caixia

doi:10.1038/s41587-021-00868-w

Cited by 234 publications

(166 citation statements)

References 22 publications

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“…Although it is a relatively new technique, prime editing has been used in many different cell lines and organisms including plants, zebrafish and Drosophila [8][9][10][11][12][13][14][15][16][17][18][19][20]. In this study we have created versatile plasmids to perform prime editing PE2, PE3 and PE-nuclease system in mammalian cells.…”

Section: Discussionmentioning

confidence: 99%

Optimized nickase- and nuclease-based prime editing in human and mouse cells

Adikusuma

Lushington

Arudkumar

et al. 2021

Preprint

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Precise genomic modification using prime editing (PE) holds enormous potential for research and clinical applications. Currently, the delivery of PE components to mammalian cell lines requires multiple plasmid vectors. To overcome this limitation, we generated all-in-one prime editing (PEA1) constructs that carry all the components required for PE, along with a selection marker. We tested these constructs (with selection) in HEK293T, K562, HeLa and mouse embryonic stem (ES) cells. We discovered that PE efficiency in HEK293T cells was much higher than previously observed, reaching up to 95% (mean 67%). The efficiency in K562 and HeLa cells, however, remained low. To improve PE efficiency in K562 and HeLa, we generated a nuclease prime editor and tested this system in these cell lines as well as mouse ES cells. PE-nuclease generated intended edits with efficiencies that were similar, and in some cases exceeded, the PE-nickase system. We also show that the nuclease prime editor can generate intended modifications in mouse fetuses with up to 100% efficiency.

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Section: Discussionmentioning

confidence: 99%

Optimized nickase- and nuclease-based prime editing in human and mouse cells

Adikusuma

Lushington

Arudkumar

et al. 2021

Preprint

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“…While our data show the promising way to confirm the cloned plasmids for CRISPR work, there are multiple opportunities to further expand the IRI-CCE utility. For example, IRI-CCE allows testing of pegRNA functionality because it is one of the critical aspects for the successful application of prime editing [53, 54].…”

Section: Discussionmentioning

confidence: 99%

IRI-CCE: A Platform for Evaluating CRISPR-based Base Editing Tools and Its Components

Shelake

Pramanik

Kim

2021

Preprint

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Rapid assessment of Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-based tools and validation of cloned CRISPR-plasmid vectors are critical aspects for the successful application of genome editing (GE) in different organisms, including plants. Also, the selection of active guide RNAs (gRNAs) is a determining factor to achieve efficient GE at the targeted locus. Incidentally, plasmid vectors capable of ectopic expression of Cas enzyme and sgRNAs in bacteria will facilitate the quick screening and reliability of cloned plasmids and the functionality of designed gRNAs. We report a platform for in vivo rapid investigation of CRISPR components in Escherichia coli (IRI-CCE), which is compatible with plant-transforming binary vectors comprising plant promoters/terminators. Besides, IRI-CCE analyses reveal distinct features of cytidine (PmCDA1, evoCDA1, APOBEC3A) and adenine (ABE8e) base editors. Finally, we show the IRI-CCE platform as a reliable and rapid method for screening functional gRNAs to achieve successful GE outcomes. Base editor-based CRISPR components expressed by promoters of different strengths led to the establishment of IRI-CCE platform for three major applications: optimization of novel CRISPR tools, validation of cloned CRISPR plasmids, and to know gRNA functionality.

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“…[27][28][29] The frontiers of precision genome editing continues to advance, for example, the improved efficiency of prime editing in plants. 30 We note that Chinese scholars, such as Xingxu Huang, Liangxue Lai, Caixia Gao, Yongping Huang, and Qunxin She, have high scholarly outputs. In addition, they have published numerous articles in the past few years.…”

Section: Global Distribution Of Scholars In Crispr Gene Editingmentioning

confidence: 97%

A Decade of CRISPR Gene Editing in China and Beyond: A Scientometric Landscape

et al. 2021

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Since its Nobel Prize-winning breakthrough in 2012, CRISPR-Cas-based gene-editing system has emerged as one of the most promising biotechnologies in decades. In this article, we present an objective and comprehensive evaluation of CRISPR-based gene-editing technologies, including base editing and prime editing, based on the bibliometric analysis of 22,902 published records. We also assessed the status of CRISPR gene-editing technologies in academia from 2010 to 2020 globally, with respect to countries, institutions, and researchers, and used text clustering methods to assess technical trends and research hotspots. Our results indicate, not surprisingly, that this is a thriving and prominent area of research. By comparing the relevance and growth of CRISPR geneediting technologies in China with other countries by several metrics, we show that the Chinese scientific community attaches considerable importance to the field of plant genome engineering, with more scholars from agricultural sectors than other sectors.

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High-efficiency prime editing with optimized, paired pegRNAs in plants

Cited by 234 publications

References 22 publications

Optimized nickase- and nuclease-based prime editing in human and mouse cells

Optimized nickase- and nuclease-based prime editing in human and mouse cells

IRI-CCE: A Platform for Evaluating CRISPR-based Base Editing Tools and Its Components

A Decade of CRISPR Gene Editing in China and Beyond: A Scientometric Landscape

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