ArticlePlant physiology2022
Insights into the molecular mechanisms of CRISPR/Cas9-mediated gene targeting at multiple loci in Arabidopsis.
Article in Plant physiology, 2022. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 12 papers.
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Who cites it
12 citing papers in PubMed.
- Occupancy-based mechanism is the chief mode of ROS1 function in preventing DNA hypermethylation.Nature plants · 2026Article
- Versatile Applications of CRISPR-Based Programmable T-DNA Integration in Plants.Plant biotechnology journal · 2025Article
- SSA-mediated selection marker gene activation enhances relative gene targeting efficiency in plants.Horticulture research · 2025Article
- Structure, regulation and assembly of the photosynthetic electron transport chain.Nature reviews. Molecular cell biology · 2025Review
- Single-stranded DNA (ssDNA) donor repair templates and CRISPR/Cas9 enable a high-frequency of targeted insertions in potato.Frontiers in genome editing · 2025Article
- Double step screening using endogenous marker improves relative gene targeting efficiency in Arabidopsis.Scientific reports · 2024Article
- aChIP is an efficient and sensitive ChIP-seq technique for economically important plant organs.Nature plants · 2024Article
- Cas12a-mediated gene targeting by sequential transformation strategy in Arabidopsis thaliana.BMC plant biology · 2024Article
- Application of multiple sgRNAs boosts efficiency of CRISPR/Cas9-mediated gene targeting in Arabidopsis.BMC biology · 2024Article
- Simple promotion of Cas9 and Cas12a expression improves gene targeting via an all-in-one strategy.Frontiers in plant science · 2024Article
- Suppression ofProceedings of the National Academy of Sciences of the United States of America · 2023Article
- Precise and heritable gene targeting in rice using a sequential transformation strategy.Cell reports methods · 2023Article
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Authors and funding
16 authors.
Funding
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Abstract
Homologous recombination-mediated gene targeting (GT) enables precise sequence knockin or sequence replacement, and thus is a powerful tool for heritable precision genome engineering. We recently established a clustered regularly interspaced short palindromic repeats/clustered regularly interspaced short palindromic repeats-associated protein 9 (CRISPR/Cas9)-mediated approach for heritable GT in Arabidopsis (Arabidopsis thaliana), but its broad utility was not tested, and the underlying molecular mechanism was unclear. Here, we achieved precise GT at 14 out of 27 tested endogenous target loci using the sequential transformation approach and obtained vector-free GT plants by backcrossing. Thus, the sequential transformation GT method provides a broadly applicable technology for precise genome manipulation. We show that our approach generates heritable GT in the egg cell or early embryo of T1 Arabidopsis plants. Analysis of imprecise GT events suggested that single-stranded transfer DNA (T-DNA)/VirD2 complexes produced during the Agrobacterium (Agrobacterium tumefaciens) transformation process may serve as the donor templates for homologous recombination-mediated repair in the GT process. This study provides new insights into the molecular mechanisms of CRISPR/Cas9-mediated GT in Arabidopsis.
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