2026-04-17T14:48:53Zhttps://recercat.cat/oai/request

oai:recercat.cat:10230/429552025-12-12T03:33:28Zcom_2072_6col_2072_452952

00925njm 22002777a 4500 dc Yang, Jae-Seong author Garriga-Canut, Mireia author Link, Nele author Carolis, Carlo author Broadbent, Katrina author Beltran-Sastre, Violeta author Serrano Pubull, Luis, 1982- author Maurer, Sebastian author 2019-11-25T08:32:22Z 2019-11-25T08:32:22Z 2018 Knowing which proteins and RNAs directly interact is essential for understanding cellular mechanisms. Unfortunately, discovering such interactions is costly and often unreliable. To overcome these limitations, we developed rec-YnH, a new yeast two and three-hybrid-based screening pipeline capable of detecting interactions within protein libraries or between protein libraries and RNA fragment pools. rec-YnH combines batch cloning and transformation with intracellular homologous recombination to generate bait–prey fusion libraries. By developing interaction selection in liquid–gels and using an ORF sequence-based readout of interactions via next-generation sequencing, we eliminate laborious plating and barcoding steps required by existing methods. We use rec-Y2H to simultaneously map interactions of protein domains and reveal novel putative interactors of PAR proteins. We further employ rec-Y2H to predict the architecture of published coprecipitated complexes. Finally, we use rec-Y3H to map interactions between multiple RNA-binding proteins and RNAs—the first time interactions between protein and RNA pools are simultaneously detected. We also acknowledge the support from the Spanish Ministry of Economy and Competitiveness (MINECO) for Juan de la Cierva-Incorporación Programme (IJCI‐2014‐22070) to J.-S.Y., L.S. (BFU2015-63571-P), and S.M. (BFU2014-54278-P and BFU2015-62550-ERC). We further acknowledge support of the Spanish Ministry of Economy and Competitiveness, “Centro de Excelencia Severo Ochoa 2013-2017”, SEV-2012-0208 and the CERCA Programme/Generalitat de Catalunya. This work was funded by the Spanish Ministry of Economy, Industry and Competitiveness (MEIC) reference MINECO PE 2013-2016 PN FEDER and the European Regional Development Fund (ERDF). All sequencing was done in the CRG Genomics Core Facility. Assay systems High-throughput screening rec-YnH enables simultaneous many-by-many detection of direct protein-protein and protein-RNA interactions