What is an RNAi screen?
Like genetic screening, RNAi screening allows for identification of genes relevant to a given pathway, structure or function via association of a mutant phenotype with gene knockdown. Like chemical screening, RNAi screening is amenable to miniaturization and automation, facilitating high-throughput studies.
What is genome wide RNAi screen?
Genome-wide RNA interference (RNAi) screening is an emerging and powerful technique for genetic screens, which can be divided into arrayed RNAi screen and pooled RNAi screen/selection based on different screening strategies.
What is RNAi and why is it important?
RNA interference (RNAi) is a post-transcriptional, highly conserved process in eukaryotes that leads to specific gene silencing through degradation of the target mRNA. This mechanism is mediated by double-stranded RNA (dsRNA) that is homologous in sequence to the silenced gene.
What can cause RNAi?
Endogenous triggers of RNAi pathway include foreign DNA or double-stranded RNA (dsRNA) of viral origin, aberrant transcripts from repetitive sequences in the genome such as transposons, and pre-microRNA (miRNA).
How does the RNAi work?
RNAi is short for “RNA interference” and it refers to a phenomenon where small pieces of RNA can shut down protein translation by binding to the messenger RNAs that code for those proteins. RNA interference is a natural process with a role in the regulation of protein synthesis and in immunity.
How are RNAi reagents used in RNAi screening?
RNAi screening relies on the availability of genome-wide or other large-scale RNAi reagent libraries, withone or more unique RNAi reagent directed againsteach target gene. The appropriate RNAi reagentlibrary for cell-based screening depends upon the celltype, approach and method of reagent delivery.7In
How is a genome wide RNAi screen used?
Genome-wide RNAi screens are most successful when they incorporate a focused search for predicted regulatory proteins. Drosophila RNAi screens have been used to identify regulators of store-operated Ca 2+ entry, a process necessary for sustained nuclear translocation of the transcription factor NFAT 7–9.
Which is an example of a siRNA screen?
In an informative example of an siRNA screen, a classical assay for cell motility was adapted to a high-throughput screening format through the reproducible use of a robotic pin to ‘wound’ confluent monolayers of MCF-10A breast epithelial cells cultured in 96-well plates.