A5086834798- CRISPR and Genetic Engineering
- Advanced biosensing and bioanalysis techniques
- Pluripotent Stem Cells Research
- Insect symbiosis and bacterial influences
- RNA Interference and Gene Delivery
- Viral Infectious Diseases and Gene Expression in Insects
- Plant Virus Research Studies
- Virus-based gene therapy research
- RNA and protein synthesis mechanisms
- Evolution and Genetic Dynamics
- Genetics and Neurodevelopmental Disorders
- Genetics, Aging, and Longevity in Model Organisms
- DNA Repair Mechanisms
- Chromosomal and Genetic Variations
- Mosquito-borne diseases and control
- HIV Research and Treatment
- Innovation and Socioeconomic Development
- Molecular Biology Techniques and Applications
Hiroshima University
2014-2023
Massachusetts Institute of Technology
2022
Genome engineering using programmable nucleases enables homologous recombination (HR)-mediated gene knock-in. However, the labour used to construct targeting vectors containing homology arms and difficulties in inducing HR some cell type organisms represent technical hurdles for application of HR-mediated knock-in technology. Here, we introduce an alternative strategy transcription activator-like effector (TALENs) clustered regularly interspaced short palindromic repeats...
Abstract The CRISPR/Cas9 system provides a powerful tool for genome editing in various model organisms, including zebrafish. establishment of targeted gene-disrupted zebrafish (knockouts) is readily achieved by CRISPR/Cas9-mediated modification. Recently, exogenous DNA integration into the via homology-independent repair was reported, but this contained mutations at junctions genomic and integrated DNA. Thus, precise modification loci remains to be achieved. Here, we describe efficient,...
Although CRISPR/Cas enables one-step gene cassette knock-in, assembling targeting vectors containing long homology arms is a laborious process for high-throughput knock-in. We recently developed the CRISPR/Cas-based precise integration into target chromosome (PITCh) system knock-in without mediated by microhomology-mediated end-joining. Here, we identified exonuclease 1 (Exo1) as an enhancer PITCh in human cells. By combining Exo1 and PITCh-directed donor vectors, achieved convenient of...
The emerging genome editing technology has enabled the creation of gene knock-in cells easily, efficiently, and rapidly, which dramatically accelerated research in field mammalian functional genomics, including humans. We recently developed a microhomology-mediated end-joining-based method, termed PITCh system, presented various examples its application. Since system only requires very short microhomologies (up to 40 bp) single-guide RNA target sites on donor vector, targeting construct can...
Selective genome editing such as gene knock-in has recently been achieved by administration of chemical enhancer or inhibitor particular DNA double-strand break (DSB) repair pathways, well overexpression pathway-specific genes. In this study, we attempt to enhance the efficiency further secure robust knock-ins, using local accumulation DSB molecules (LoAD) system. We identify CtIP a strong microhomology-mediated end-joining (MMEJ) genetic screening, and show knock-in-enhancing effect...
Single‐cell cloning is an essential technique for establishing genome‐edited cell clones mediated by programmable nucleases such as CRISPR ‐Cas9. However, residual genome‐editing activity after single‐cell may cause heterogeneity in the clonal cells. Previous studies showed efficient mutagenesis and rapid degradation of ‐Cas9 components cultured cells introducing Cas9 ribonucleoproteins ( RNP s). In this study, we investigated how timing ‐transfected affected resultant clones. We carried out...
Abstract Efficiency and accuracy are paramount in genome editing. While CRISPR-Cas nucleases efficient at editing target genes, their is limited because following DNA cleavage by Cas proteins, error-prone repair mechanisms introduce random mutations. Improving the of reducing repairs using DNA- or RNA-based templates can compromise efficiency. To simultaneously improve both efficiency accuracy, we created a frameshifting genome-editing technology fusing Cas9 with polymerases. These Frame...
Abstract Highly efficient gene knock-out and knock-in have been achieved by harnessing CRISPR-Cas9 its advanced technologies such as Prime Editor. In addition, various bioinformatics resources become available to quantify qualify the efficiency accuracy of CRISPR edits, which significantly increased user-friendliness general next-generation sequencing (NGS) analysis in context genome editing. However, there is no specialized integrated software for investigating preference genomic involved...
Abstract Highly efficient gene knock-out and knock-in have been achieved by harnessing CRISPR-Cas9 its advanced technologies such asPrime Editor. In addition, various bioinformatics resources become available to quantify qualify the efficiency accuracy of CRISPR edits, which significantly increased user-friendliness general next-generation sequencing (NGS) analysis in context genome editing. However, there is no specialized integrated software for investigating preference genomic involved...