Baculoviral delivery of CRISPR/Cas9 facilitates efficient genome editing in human cells
| Autor: | Jasmin B. Post, My Anh Truong, Susanne M.A. Lens, Ingrid Verlaan-Klink, Hugo J. Snippert, Arne J. Bramer, Sanne Hindriksen, Michael A. Hadders, Martijn J.M. Vromans |
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| Jazyk: | angličtina |
| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
lcsh:Medicine Artificial Gene Amplification and Extension Baculoviruses Synthetic Genome Editing Polymerase Chain Reaction Genome Genome Engineering Gene Knockout Techniques Transduction (genetics) Genome editing CRISPR Cell Cycle and Cell Division lcsh:Science Gene Editing Centromeres Genetics Multidisciplinary Chromosome Biology Intracellular Signaling Peptides and Proteins Crispr Cell Processes Viruses Engineering and Technology Synthetic Biology Baculoviridae Research Article Biotechnology Chromosome Structure and Function Mitosis Bioengineering Computational biology Protein Serine-Threonine Kinases Biology Research and Analysis Methods Chromosomes Viral vector Homology directed repair Gene Delivery 03 medical and health sciences Cell Line Tumor Gene Expression and Vector Techniques Journal Article Humans Point Mutation Molecular Biology Techniques Molecular Biology Molecular Biology Assays and Analysis Techniques CRISPR interference Genome Human Cas9 lcsh:R Organisms Biology and Life Sciences Cell Biology Synthetic Genomics 030104 developmental biology Synthetic Bioengineering Mutation lcsh:Q CRISPR-Cas Systems DNA viruses Cloning |
| Zdroj: | PLoS ONE [E], 12(6). Public Library of Science PLoS ONE PLoS ONE, Vol 12, Iss 6, p e0179514 (2017) |
| ISSN: | 1932-6203 |
| Popis: | The CRISPR/Cas9 system is a highly effective tool for genome editing. Key to robust genome editing is the efficient delivery of the CRISPR/Cas9 machinery. Viral delivery systems are efficient vehicles for the transduction of foreign genes but commonly used viral vectors suffer from a limited capacity in the genetic information they can carry. Baculovirus however is capable of carrying large exogenous DNA fragments. Here we investigate the use of baculoviral vectors as a delivery vehicle for CRISPR/Cas9 based genome-editing tools. We demonstrate transduction of a panel of cell lines with Cas9 and an sgRNA sequence, which results in efficient knockout of all four targeted subunits of the chromosomal passenger complex (CPC). We further show that introduction of a homology directed repair template into the same CRISPR/Cas9 baculovirus facilitates introduction of specific point mutations and endogenous gene tags. Tagging of the CPC recruitment factor Haspin with the fluorescent reporter YFP allowed us to study its native localization as well as recruitment to the cohesin subunit Pds5B. |
| Databáze: | OpenAIRE |
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