Crispr/cas9 Gene Disruption
Crispr/cas9 Gene Disruption - Related Videos
04:22
A Technique for Gene Editing in Natural Killer Cells Using CRISPR Cas9
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2025
This video demonstrates a technique for Cas9 ribonucleoprotein-mediated genetic modification of primary natural killer (NK) cells. A Cas9 ribonucleoprotein, consisting of a Cas9 endonuclease bound to a guide RNA (gRNA) formed by base pairing a CRISPR RNA (crRNA) and a trans-activating crRNA (tracrRNA), is introduced into primary natural killer cells via electroporation. The ribonucleoprotein targets and cleaves the host DNA at the target site, leading to gene knockout via modification of the...
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.
11:37
QTL Mapping and CRISPR/Cas9 Editing to Identify a Drug Resistance Gene in Toxoplasma gondii
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Cited by 5 •
2017
Details are presented on how QTL mapping with a whole genome sequence based genetic map can be used to identify a drug resistance gene in Toxoplasma gondii and how this can be verified with the CRISPR/Cas9 system that efficiently edits a genomic target, in this case the drug resistance gene.
05:12
All-in-One CRISPR Genome Editing: A Method for Homology Directed Repair-Based Gene Knock-In in Cultured Cells Using CRISPR-Cas9 System
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2025
In this video, we demonstrate all-in-one CRISPR-Cas9 based genome editing in cultured cells where Cas9 and sgRNA are provided as a single plasmid construct to the cells. The CRISPR-Cas9 system and desired gene to be inserted was introduced in cells through electroporation technique to facilitate successful gene editing.
FAQs
The PAM acts as a required neighboring sequence for target recognition: the guide RNA pairs with the complementary DNA sequence, but Cas9 cuts only where that target is positioned next to an appropriate PAM. This requirement constrains which genomic sites can be selected, making PAM availability an important consideration during guide design and experimental planning.
After Cas9 makes the double-strand break, error-prone non-homologous end joining can reconnect the DNA while introducing insertions or deletions. If these changes interrupt the gene’s coding sequence, they can reduce or eliminate its function. The resulting disruption therefore depends on how repair-generated sequence changes affect the targeted coding region.
Guide design helps focus Cas9 activity on the intended gene region, whereas verification tests whether the expected on-target sequence change actually occurred and helps identify unintended sequence changes. Using both steps strengthens interpretation: an observed phenotype can be linked more confidently to disruption of the selected gene rather than to an unrecognized editing event.
A basic workflow begins by selecting a guide RNA complementary to a target sequence beside a PAM, then using the guide-Cas9 system to create the double-strand break. The edited locus is subsequently checked for the intended sequence change. This sequence of design, cleavage, repair, and verification connects the molecular intervention to a usable genetic result.
In functional screens, disrupting genes provides a way to test which genes contribute to a cellular trait or phenotype. Researchers can compare the consequences of gene interruption across targets, then use selected results to validate gene-phenotype relationships. The approach therefore links targeted genome changes with systematic investigation of gene function.
Researchers apply this strategy when they need cellular or animal models in which a gene’s function has been reduced or eliminated. Such models can help examine how a genetic disruption relates to disease-associated phenotypes and support testing of gene-phenotype relationships. In genetics, they extend gene editing from individual loci to model-based biological investigation.