ArticleMolecular biology of the cell2021
One-step multiplex toolkit for efficient generation of conditional gene silencing human cell lines.
Article in Molecular biology of the cell, 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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10 citing papers in PubMed, 13 citations in OpenAlex.
- Plasticity of mitotic cyclins in promoting the G2-M transition.The Journal of cell biology · 2025Article
- Kinesins regulate the heterogeneity in centrosome clustering after whole-genome duplication.Life science alliance · 2024Article
- Coregulation of NDC80 Complex Subunits Determines the Fidelity of the Spindle-Assembly Checkpoint and Mitosis.Molecular cancer research : MCR · 2024Article
- HiHo-AID2: boosting homozygous knock-in efficiency enables robust generation of human auxin-inducible degron cells.Genome biology · 2024Article
- NHE7 upregulation potentiates the uptake of small extracellular vesicles by enhancing maturation of macropinosomes in hepatocellular carcinoma.Cancer communications (London, England) · 2024Article
- BCL-XL regulates the timing of mitotic apoptosis independently of BCL2 and MCL1 compensation.Cell death & disease · 2024Article
- A robust dual gene ON-OFF toggle directed by two independent promoter-degron pairs.Journal of cell science · 2023Article
- Cyclin A-CDK1 suppresses the expression of the CDK1 activator CDC25A to safeguard timely mitotic entry.The Journal of biological chemistry · 2023Article
- MARCH5 regulates mitotic apoptosis through MCL1-dependent and independent mechanisms.Cell death and differentiation · 2023Article
- The human EF1a promoter does not provide expression of the transgene in mice.Transgenic research · 2022Article
Corrections and comments
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Authors and funding
4 authors at 2 institutions in 3 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Loss-of-function analysis is one of the major arsenals we have for understanding gene functions in mammalian cells. For analysis of essential genes, the major challenge is to develop simple methodologies for tight and rapid inducible gene inactivation. One approach involves CRISPR-Cas9-mediated disruption of the endogenous locus in conjunction with the expression of a rescue construct, which can subsequently be turned off to produce a gene inactivation effect. Here we describe the development of a set of Sleeping Beauty transposon-based vectors for expressing auxin-inducible degron (AID)-tagged genes under the regulation of a tetracycline-controlled promoter. The dual transcriptional and degron-mediated post-translational regulation allows rapid and tight silencing of protein expression in mammalian cells. We demonstrated that both non-essential and essential genes could be targeted in human cell lines using a one-step transfection method. Moreover, multiple genes could be simultaneously or sequentially targeted, allowing inducible inactivation of multiple genes. These resources enable highly efficient generation of conditional gene silencing cell lines to facilitate functional studies of essential genes.
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