Crispr-cas9 Genome Engineering
Crispr-cas9 Genome Engineering - Related Videos
08:20
Genome Engineering of Primary Human B Cells Using CRISPR/Cas9
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Cited by 13 •
2020
Here we provide a detailed, step-by-step protocol for CRISPR/Cas9-based genome engineering of primary human B cells for gene knockout (KO) and knock-in (KI) to study biological functions of genes in B cells and the development of B-cell therapeutics.
06:12
Production of Genetically Engineered Golden Syrian Hamsters by Pronuclear Injection of the CRISPR/Cas9 Complex
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Cited by 24 •
2018
Pronuclear (PN) injection of the clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated protein-9 nuclease (CRISPR/Cas9) system is a highly efficient method for producing genetically engineered golden Syrian hamsters. Herein, we describe the detailed PN injection protocol for the production of gene knockout hamsters with the CRISPR/Cas9 system.
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.
01:28
CRISPR/Cas9 Genome Editing
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2025
The CRISPR-Cas system serves as a bacterial defense mechanism against invading genetic elements such as viruses and plasmids, forming the foundation for its adaptation as a powerful genome-editing tool. Originally discovered in prokaryotes, this system has been repurposed to revolutionize genetic engineering across a wide range of organisms, including plants, animals, and humans. The core component, Cas9, is an endonuclease derived from Streptococcus pyogenes, capable of introducing...
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 protospacer adjacent motif, or PAM, provides a required DNA landmark next to the sequence recognized by the guide RNA. Cas9 acts at a complementary target only when this neighboring motif is present, so PAM availability influences which genomic sites can be selected. This requirement helps determine the possible targets for studying host or pathogen genes.
After Cas9 creates a double-strand break, the cell can repair the lesion through error-prone or template-directed pathways. Error-prone repair can disrupt a gene, whereas template-directed repair can introduce a defined sequence change when suitable template information is available. The selected repair outcome therefore determines whether an experiment primarily creates gene disruption or a planned mutation.
Targeting depends on complementarity between the guide RNA and the selected DNA sequence, together with the presence of a nearby PAM. These two features work together to position Cas9 at the intended locus. In genome-engineering experiments, selecting a target with both characteristics is central to connecting a DNA change with the gene function being investigated.
The same editing framework can be directed toward host genes or pathogen genes, allowing researchers to examine different sides of an infection process. Altering host genes can reveal contributions to immune signaling or viral entry, while altering pathogen genes can expose mechanisms of microbial survival. Comparing these outcomes helps assign functions within host-pathogen interactions.
A basic workflow identifies a DNA sequence of interest, selects a complementary guide RNA target next to a PAM, and uses Cas9 to create a double-strand break at that site. Cellular repair then produces either an error-prone disruption or a template-directed change. The resulting genetic model can be used to study the function of the edited gene.
This approach is useful when researchers need to test whether a particular host or pathogen gene contributes to infection or immune responses. Editing can support investigations of immune signaling, viral entry, and microbial survival, while also enabling disease modeling and target validation. These results can identify genetic processes relevant to potential therapeutic development.
Defined disruptions or mutations create models in which the contribution of a selected gene can be examined directly. In immunology and infection studies, such models can connect genetic changes with altered immune signaling, viral entry, or microbial survival. This evidence supports target validation by showing whether a gene or pathway is relevant to the disease process.
The repair response can generate either disruptive changes or defined mutations, giving researchers more than one way to investigate infection biology. Disruptions can test whether a gene is required, while defined mutations can examine the effect of a particular sequence change. Applying these outcomes to host or pathogen genes helps resolve mechanisms of infection and immunity.