ArticleScientific reports2021
Construction of adenovirus vectors simultaneously expressing four multiplex, double-nicking guide RNAs of CRISPR/Cas9 and in vivo genome editing.
Article in Scientific reports, 2021. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 10 papers.
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Who cites it
10 citing papers in PubMed, 23 citations in OpenAlex.
- CRISPR-Cas9: Genome Engineering and Future Vaccine Applications.Molecular biotechnology · 2026Review
- Disruption of Human Papillomavirus 16 E6/E7 Genes Using All-in-One Adenovirus Vectors Expressing Eight Double-Nicking Guide RNAs.International journal of molecular sciences · 2025Article
- The Role of CRISPR/Cas9 in Revolutionizing Duchenne's Muscular Dystrophy Treatment: Opportunities and Obstacles.Global medical genetics · 2024Review
- Construction and Stability of All-in-One Adenovirus Vectors Simultaneously Expressing Four and Eight Multiplex Guide RNAs and Cas9 Nickase.International journal of molecular sciences · 2024Article
- Extracellular matrix re-normalization to improve cold tumor penetration by oncolytic viruses.Frontiers in immunology · 2024Review
- Intraplacental injection of AAV9-CMV-iCre results in the widespread transduction of multiple organs in double-reporter mouse embryos.Experimental animals · 2023Article
- Gene therapy for alopecia in type II rickets model rats using vitamin D receptor-expressing adenovirus vector.Scientific reports · 2023Article
- Article
- Short term but highly efficient Cas9 expression mediated by excisional system using adenovirus vector and Cre.Scientific reports · 2021Article
- Adenovirus Vectors Expressing Eight Multiplex Guide RNAs of CRISPR/Cas9 Efficiently Disrupted Diverse Hepatitis B Virus Gene Derived from Heterogeneous Patient.International journal of molecular sciences · 2021Article
Corrections and comments
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Authors and funding
7 authors at 3 institutions in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Simultaneous expression of multiplex guide RNAs (gRNAs) is valuable for knockout of multiple genes and also for effective disruption of a gene by introducing multiple deletions. We developed a method of Tetraplex-guide Tandem for construction of cosmids containing four and eight multiplex gRNA-expressing units in one step utilizing lambda in vitro packaging. Using this method, we produced an adenovirus vector (AdV) containing four multiplex-gRNA units for two double-nicking sets. Unexpectedly, the AdV could stably be amplified to the scale sufficient for animal experiments with no detectable lack of the multiplex units. When the AdV containing gRNAs targeting the H2-Aa gene and an AdV expressing Cas9 nickase were mixed and doubly infected to mouse embryonic fibroblast cells, deletions were observed in more than 80% of the target gene even using double-nicking strategy. Indels were also detected in about 20% of the target gene at two sites in newborn mouse liver cells by intravenous injection. Interestingly, when one double-nicking site was disrupted, the other was simultaneously disrupted, implying that two genes in the same cell may simultaneously be disrupted in the AdV system. The AdVs expressing four multiplex gRNAs could offer simultaneous knockout of four genes or two genes by double-nicking cleavages with low off-target effect.
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