Native gel analysis for mammalian microRNPs assembled from pre-microRNAs

MicroRNAs (miRNAs) are an important class of small RNAs that regulate gene expression posttranscriptionally through the microRNP (miRNP)/RNA-induced silencing complex (RISC). The core component of miRNPs is an Argonuate protein that directly binds to a miRNA. In mammals, most miRNPs are assembled through the miRNA loading complex (miRLC), which is composed of Dicer, Ago, and TRBP. miRLC processes miRNA precursors (pre-miRNAs) into miRNA duplexes and loads miRNA duplexes to Ago. Here, we describe a native gel analysis system for detecting miRNPs assembled with pre-miRNAs from mammalian lysates that ectopically express Ago2. The methods presented here provide a powerful tool for further dissecting miRNP assembly pathways in mammals.