What is shRNA transfection?

What is shRNA transfection?

What is shRNA transfection?

siRNA Transfection. Small interfering RNA (siRNA) is a 20-28 nucleotide double stranded RNA (dsRNA) molecule often referred to as “silencing RNA” . siRNA is used in the gene silencing (or RNA Interference, RNAi) technique to suppress gene expression.

What is retroviral transfection?

Retroviruses have the ability to transform their single-stranded RNA genome into a double-stranded DNA molecule that stably integrates into the genome of dividing target cells. Retroviral transduction has been widely used for cancer and stem cell research.

What does shRNA target?

A short hairpin RNA or small hairpin RNA (shRNA/Hairpin Vector) is an artificial RNA molecule with a tight hairpin turn that can be used to silence target gene expression via RNA interference (RNAi). Expression of shRNA in cells is typically accomplished by delivery of plasmids or through viral or bacterial vectors.

How are retroviral vectors made?

A retroviral vector is produced by inserting the transgene in place of part of the viral genome, and a preparation of infectious viral particles is produced by introducing the recombinant virus into tissue culture cells.

How are shRNAs transfected in mammalian cells?

Like siRNAs, shRNAs may be transfected as plasmid vectors encoding shRNAs transcribed by RNA pol III or modified pol II promoters, but can also be delivered into mammalian cells through infection of the cell with virally produced vectors.

Is it possible to transfect primary cells with shRNA with adenovirus?

Lentivirus and adenovirus have made it possible for cells that are refractory to transfection such as primary cells to become permissive to shRNA. However, as these methods are based on a viral backbone, each method harbors inherent dangers which would limit their use to in vitro studies.

Who prepared the retrovirus transfection protocol?

Retrovirus Transfection Protocol Prepared by: Berggren, Travis [email protected] Date Submitted April 24, 2012 Submitted by Berggren, Travis [email protected] Adapted from Salk Stem Cell Core in-house protocols Contributor(s) Lutz, Margaret. Modesto, Veronica. Panopoulos, Athanasia. Affiliation The Salk Institute Introduction:

What happens to the shRNA after it is transcribed?

Following transcription, the shRNA sequence is exported to the cytosol where it is recognized by an endogenous enzyme, Dicer, which processes the shRNA into the siRNA duplexes (seeChapter 7).