Related Experiment Video
Updated: Mar 24, 2026

Protein Extract Preparation and Co-immunoprecipitation from Caenorhabditis elegans
Published on: May 23, 2020
Loading of Argonaute Protein with Small Duplex RNA in Cellular Extracts
1Department of Biochemistry and Molecular Biology, School of Medicine, Southern Illinois University, Carbondale, IL, 62901, USA. ktgagnon@siu.edu.
Abstract:
Argonaute (Ago) proteins are the minimum core proteins required for executing RNA interference (RNAi) mechanisms of gene regulation. For Ago proteins to regulate gene expression through RNAi they must be loaded, or "programmed," with a single strand of small RNA. Natural small RNAs are typically double-stranded duplexes that require additional factors for efficient and specific loading into Ago proteins. Here, a protocol is described for investigating RNAi programming through loading of human Ago2 using radiolabeled small interfering RNA (siRNA) and HeLa cell extracts. This protocol provides an Ago loading assay to study RNAi programming when starting with crude or partially purified cell extracts. The Ago loading assay should prove useful for studying other Ago proteins using a variety of mammalian cell extracts.
Related Concept Videos
RNA Interference
This process occurs naturally in cells, often through the activity of genomically-encoded microRNAs. Researchers can take advantage of this mechanism by introducing synthetic RNAs to deactivate specific genes for research or therapeutic purposes. For example, RNAi could be used...
siRNA - Small Interfering RNAs
In the cytoplasm, siRNA is processed from a double-stranded RNA, which comes from either endogenous DNA transcription or exogenous sources like a virus. This double-stranded RNA is then cleaved by the...
Experimental RNAi
piRNA - Piwi-interacting RNAs
Conservation of Protein Domains Over Different Proteins
A limited set of protein domains often duplicate and recombine during evolution. These domains can be organized in different combinations to...

