Dd-cas9 Destabilization
Dd-cas9 Destabilization - Related Videos
01:45
Destabilization of Microtubules
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2023
The destabilization of microtubules can occur during different stages of the microtubule lifecycle, such as nucleation or elongation. It can take place at either end of the microtubule or in the microtubule lattices as a whole. The lifespan of individual microtubules within a cell varies according to the cell type and stage of the cell cycle. During interphase, the lifespan of the microtubule is about 30 minutes, while during cell division, it is about 15 minutes. In axonal microtubules of...
01:10
Drugs that Destabilize Microtubules
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2023
Microtubules are dynamic structures and can be regulated by microtubule targeting agents (MTAs). Microtubule destabilizing drugs are a class of MTAs that destabilize and prevent microtubules' polymerization. Both natural and synthetic chemicals can be found under this class of drugs. Vincristine and vinblastine, two vinca alkaloids, and colchicine were among the first to be discovered. These drugs can affect cells in various ways, either by inducing a change in cell morphology, preventing...
08:05
A Prediction Error-driven Retrieval Procedure for Destabilizing and Rewriting Maladaptive Reward Memories in Hazardous Drinkers
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Cited by 24 •
2018
This manuscript describes a memory retrieval procedure for destabilizing robust reward memories and rewriting them with counterconditioning prior to their reconsolidation.
02:30
A CRISPR-Cas9 Technique for Gene Editing in T Cells
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2025
This video demonstrates an assay for performing gene editing in human T cells using the CRISPR-Cas9 technology. A mixture of primary CD4+ and CD8+ T cells is combined with a CRISPR-Cas9 ribonucleoprotein complex, targeting specific genes for knockout. Upon electroporation, the sgRNA guides Cas9 to the target DNA sequence, creating precise cuts. These cuts are then repaired by the cell's non-homologous end-joining mechanism, leading to gene knockout.
02:55
Genetically Modifying CAR T Cells Using a CRISPR-Cas9 System
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2025
The video outlines a process for creating genetically modified CAR T cells through the CRISPR-Cas9 System. Infecting T cells with CRISPR and CAR lentiviruses results in modifications to the target gene and the synthesis of a chimeric antigen receptor or CAR, ultimately leading to the formation of genetically modified CAR T cells.
FAQs
The destabilization domain marks the Cas9 fusion for proteasomal degradation when a stabilizing ligand is absent. Proteasomes are cellular protein-degradation complexes, so this process lowers the amount of active fusion protein available for genome editing. The ligand Shield-1 protects the fusion from that degradation, linking chemical treatment to the presence and activity of Cas9.
Shield-1 serves as the conditional trigger for DD-Cas9 activity. Before ligand addition, degradation keeps the fusion protein restricted; after addition, protection allows Cas9 to accumulate and support editing. This arrangement lets an experimenter select when editing becomes possible, making the timing of a genetic perturbation an explicit experimental variable rather than an uncontrolled consequence of protein expression.
Restricting Cas9 availability narrows the period during which the protein can modify genomic targets. That timing helps researchers relate a resulting genetic change to a defined experimental window, which is valuable when studying gene function. Limiting activity may also reduce unintended editing by avoiding prolonged exposure to an active genome-editing protein, although the strategy does not eliminate the need for careful experimental design.
A continuously available Cas9 system does not use ligand-dependent stabilization to regulate the protein's abundance. DD-Cas9 adds a control layer at the protein level: degradation dominates under normal conditions, while Shield-1 permits stabilization. The distinction matters when researchers need to coordinate genome editing with a particular stage or treatment and to reduce the duration of Cas9 exposure.
A typical workflow begins with a genetic construct encoding Cas9 fused to a destabilization domain. The system is maintained without the stabilizing ligand so the fusion remains subject to degradation, then Shield-1 is added to permit protein stabilization and editing. Researchers can therefore align ligand treatment with the intended perturbation window and assess gene-function consequences within that controlled design.
DD-Cas9 is particularly useful when the timing of a genome perturbation matters to the biological question. In genetics, researchers can use ligand-controlled Cas9 activity to introduce a defined intervention and then examine its functional consequences. The approach also supports experimental designs that seek greater temporal control and potentially less unintended editing than prolonged Cas9 activity would provide.