Ribonucleoprotein Delivery
Ribonucleoprotein Delivery - Related Videos
09:23
Delivery of the Cas9/sgRNA Ribonucleoprotein Complex in Immortalized and Primary Cells via Virus-like Particles ("Nanoblades")
0 Views •
Cited by 6 •
2021
We have developed a simple and inexpensive protocol to load Cas9/single-guide RNA (sgRNA) ribonucleoprotein complexes within virus-like particles. These particles, called "Nanoblades", allow efficient delivery of the Cas9/sgRNA complex in immortalized and primary cells as well as in vivo.
09:35
Purification and Visualization of Influenza A Viral Ribonucleoprotein Complexes
0 Views •
Cited by 8 •
2009
The genome of the influenza A virus consists of eight separate complexes of RNA and proteins, termed viral ribonucleoprotein complexes (vRNPs). This paper describes the glycerol gradient purification and transmission electron microscopy visualization of influenza A vRNPs.
09:45
Electroporation-Mediated Delivery of Cas9 Ribonucleoproteins and mRNA into Freshly Isolated Primary Mouse Hepatocytes
0 Views •
Cited by 3 •
2022
This protocol describes techniques for isolating primary mouse hepatocytes from the liver and electroporating CRISPR-Cas9 as ribonucleoproteins and mRNA to disrupt a therapeutic target gene associated with an inherited metabolic disease of the liver. The methods described result in high viability and high levels of gene modification after electroporation.
11:20
Affinity Purification of Influenza Virus Ribonucleoprotein Complexes from the Chromatin of Infected Cells
0 Views •
Cited by 3 •
2012
Influenza viruses replicate their RNA genome in association with host-cell chromatin. Here, we present a method to purify intact viral ribonucleoprotein complexes from the chromatin of infected cells. Purified viral complexes can be analyzed by both Western blot and primer extension of protein and RNA content, respectively.
13:34
Method for the Isolation and Identification of mRNAs, microRNAs and Protein Components of Ribonucleoprotein Complexes from Cell Extracts using RIP-Chip
0 Views •
Cited by 19 •
2012
A step by step protocol to isolating and identifying RNA associated complexes through RIP-Chip.
FAQs
The guide RNA provides sequence recognition by pairing with a complementary nucleic acid sequence. This interaction positions the programmable nuclease, such as Cas9, at the selected target rather than relying on broad, nonspecific activity. Target recognition therefore connects the chosen guide sequence to the location where a targeted break or change in gene activity may occur.
The RNA component supplies the targeting information, while the protein component performs the programmable molecular action. In a Cas9-based complex, the guide RNA identifies the complementary sequence and Cas9 can create a targeted break. Separating these roles within one complex allows sequence selection and nuclease activity to function together at a defined genomic site.
Transient activity limits how long the editing components remain active inside a cell. After the guide RNA and nuclease act, the components are degraded rather than continuing to be produced from a DNA-based expression system. This shorter active period is relevant when researchers want controlled manipulation and wish to reduce prolonged component activity.
Ribonucleoprotein delivery introduces active RNA-protein complexes directly, whereas DNA-based expression systems provide genetic instructions for producing components within the cell. Direct action can produce a rapid effect and avoids the continued expression associated with those instructions. The distinction matters in experiments that prioritize temporary control over sustained production of editing components.
A typical conceptual workflow pairs a guide RNA with a programmable nuclease, introduces the assembled complex into cells, and allows the guide to direct the nuclease toward its complementary sequence. The resulting activity may create a targeted break or alter gene activity. As the components are degraded, the experiment proceeds with a transient manipulation rather than ongoing expression.
This approach supports experiments that require controlled changes in cellular genetic activity. In biology, researchers can apply it to gene editing, functional genomics, disease modeling, and investigations of cellular pathways. The resulting targeted break or altered gene activity can help connect a selected sequence or gene function with observed cellular effects.