| First Author | Li X | Year | 2019 |
| Journal | bioRxiv | Mgi Jnum | J:283893 |
| Mgi Id | MGI:6390566 | Doi | 10.1101/820548 |
| Citation | Li X, et al. (2019) High-resolution in vivo identification of miRNA targets by Halo-Enhanced Ago2 Pulldown. bioRxiv |
| abstractText | The identification of miRNA targets by Ago2 crosslink-ing-immunoprecipitation (CLIP) methods has provided major insights into the biology of this important class of non-coding RNAs. However, these methods are technically challenging and not easily translated to an in vivo setting. To overcome these limitations and to facilitate the inves-tigation of miRNA functions in mice, we have developed a method (HEAP: for Halo-Enhanced Ago2 Pulldown) to map miRNA-mRNA binding sites. This method is based on a novel genetically engineered mouse harboring a condi-tional, Cre-regulated, Halo-Ago2 allele expressed from the endogenous Ago2 locus. By using a resin conjugated to the HaloTag ligand, Ago2-miRNA-mRNA complexes can be efficiently purified from cells and tissues expressing the endogenous Halo-Ago2 allele. We demonstrate the repro-ducibility and sensitivity of this method in mouse embry-onic stem cells, in developing embryos, in adult tissues and in autochthonous mouse models of human brain and lung cancers. The tools and the datasets we have generated will serve as a valuable resource to the scientific community and will facilitate the characterization of miRNA functions under physiological and pathological conditions. |
