Cas9-grna Complex
Cas9-grna Complex - 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...
09:10
Direct Gene Knock-out of Axolotl Spinal Cord Neural Stem Cells via Electroporation of CAS9 Protein-gRNA Complexes
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Cited by 6 •
2019
Presented here is a protocol to perform time- and space-restricted gene knock-out in axolotl spinal cords by injecting CAS9-gRNA complex into the spinal cord central canal followed by electroporation.
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.
02:15
Nanoblade-Based Delivery of Nucleic Acid Cargo: A Technique to Deliver Cas9-sgRNA Complex to Target Cells via Virus-Like Particles
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2025
This video demonstrates a technique for nanoblade-based delivery of Cas9-sgRNA ribonucleoprotein complex inside target cells for genome editing. Nanoblades are virus-like particles devoid of the ability to multiply and infect neighboring cells, therefore are very useful for rapid and dose-dependent transportation of bio- and nanomaterials.
10:32
A Universal Protocol for Large-scale gRNA Library Production from any DNA Source
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Cited by 2 •
2017
Methods for generating large-scale gRNA libraries should be simple, efficient and cost-effective. We describe a protocol for the production of gRNA libraries based on enzymatic digestion of target DNA. This method, CORALINA (comprehensive gRNA library generation through controlled nuclease activity) presents an alternative to costly custom oligonucleotide synthesis.
FAQs
Recognition of the correct PAM helps position the Cas9-gRNA complex at an eligible genomic site. The guide RNA must match the complementary target sequence, while the PAM provides the adjacent sequence context needed for targeting. Consequently, a potentially matching sequence without the required neighboring PAM is not equivalent to a complete target for this editing system.
Once the target is engaged, Cas9 unwinds the DNA and produces a double-strand break. The cell’s subsequent repair route determines the editing result: error-prone repair can disrupt the gene, whereas template-directed repair can introduce a designed sequence. Thus, the nuclease cut alone does not specify whether the final outcome is a knockout or a precise modification.
Targeting accuracy matters because the intended genetic change must be distinguished from activity at other genomic locations. Off-target activity therefore remains a central research consideration when interpreting results from Cas9-gRNA experiments. This issue is especially relevant when investigators attribute a cellular or disease-model phenotype to the modification at the selected target sequence.
Gene knockout studies exploit the error-prone repair response after Cas9 cleavage. Repair can introduce changes that disrupt the targeted gene, allowing investigators to examine the consequences of losing its function. In genetics, this approach supports functional studies because observed phenotypes can be compared with the presence or absence of activity from the modified gene.
Precise genome editing depends on a different repair outcome: template-directed repair can incorporate a designed sequence after the Cas9-generated break. This makes the complex useful when researchers want to test a specific sequence change rather than simply interrupt a gene. The resulting edits can support disease modeling by reproducing or examining defined genetic alterations.
Cas9-gRNA complexes are relevant to therapeutic development because they offer a way to direct a nuclease toward a selected genomic sequence and alter DNA through cellular repair. However, outcomes depend on the repair pathway, and off-target activity can complicate confidence that only intended genomic changes occurred. These considerations keep therapeutic use an important area of ongoing research.