Volume 5, Issue 4, Pages 312-314 (October 2017) Isolation versus Enrichment: dCLIP Enables Stringent Profiling of RNA-Binding Sites  Eric L. Van Nostrand  Cell Systems  Volume 5, Issue 4, Pages 312-314 (October 2017) DOI: 10.1016/j.cels.2017.10.007 Copyright © 2017 Elsevier Inc. Terms and Conditions

Figure 1 Identification of RNA Targets with dCLIP Top: In standard RNA immunoprecipitation (RIP) or crosslinking and immunoprecipitation (CLIP), a protein of interest (green) is immunoprecipitated (with antibodies targeting either endogenous proteins or transgenic peptide tags). However, the need to preserve antibody-antigen interactions limits the stringency of wash conditions, which can lead to pulldown of other complex members (orange) and low levels of all proteins as background (other colors). Bottom: In denaturing CLIP (dCLIP), E. coli biotin holoenzyme synthetase (BirA) adds biotin specifically to a BioTag cassette added to the protein of interest. The high affinity of streptavidin for biotin enables high stringency wash conditions, leaving only the desired protein (along with a small number of endogenous proteins that are biotinylated by BirA [blue]). Cell Systems 2017 5, 312-314DOI: (10.1016/j.cels.2017.10.007) Copyright © 2017 Elsevier Inc. Terms and Conditions