biostudies-literatureUnknown8(2)Hanelt TNRNA binding proteins (RBPs) are crucial regulators of gene expression and critically depend on the specific recognition of their target RNAs. Accordingly, a selection of methods to analyze RBP specificities has been developed, including protein-RNA crosslinking and sequencing (CLIP) and in vitro selection methods such as SELEX, RNA compete or RNA bind-n-seq. However, limitations like the availability for purified recombinant proteins and custom microarray platforms (RNAcompete) or extensive sequencing depth and sophisticated bioinformatic data processing (CLIP) may limit a broader implementation of these methods. Here, we present an RNA bind-n-seq method that uses short random RNA pools and enables multiple rounds of selection. This results in strong motif enrichment with low positional vaLife science alliancee202402782https://www.ebi.ac.uk/biostudies/studies/S-EPMC11612968biostudies-literatureEndo-bind-n-seq: identifying RNA motifs of RNA binding proteins isolated from endogenous sources.PMC11612968Treiber TLehmann GReichelt RGrohmann DSchmid GZambelli FPavesi GTreiber NEichner NRothmeier THanelt TNMeister GfalseEndo-bind-n-seq: identifying RNA motifs of RNA binding proteins isolated from endogenous sources.RNA binding proteins (RBPs) are crucial regulators of gene expression and critically depend on the specific recognition of their target RNAs. Accordingly, a selection of methods to analyze RBP specificities has been developed, including protein-RNA crosslinking and sequencing (CLIP) and in vitro selection methods such as SELEX, RNA compete or RNA bind-n-seq. However, limitations like the availability for purified recombinant proteins and custom microarray platforms (RNAcompete) or extensive sequencing depth and sophisticated bioinformatic data processing (CLIP) may limit a broader implementation of these methods. Here, we present an RNA bind-n-seq method that uses short random RNA pools and enables multiple rounds of selection. This results in strong motif enrichment with low positional va2025-01-01T00:00:00Z2025 Feb2025-04-04T00:23:17.6Z2025-04-04T00:23:17.6ZS-EPMC116129683962262110.26508/lsa.202402782