| Summary: | Genome-wide physical protein–protein interaction (PPI) mapping remains a major challenge for current technologies. Here, we reported a high-efficiency BiFC-seq method, yeast-enhanced green fluorescent protein-based bimolecular fluorescence complementation (yEGFP-BiFC) coupled with next-generation DNA sequencing, for interactome mapping. We first applied yEGFP-BiFC method to systematically investigate an intraviral network of the Ebola virus. Two-thirds (9/14) of known interactions of EBOV were recaptured, and five novel interactions were discovered. Next, we used the BiFC-seq method to map the interactome of the tumor protein p53. We identified 97 interactors of p53, more than three-quarters of which were novel. Furthermore, in a more complex background, we screened potential interactors by pooling two BiFC libraries together and revealed a network of 229 interactions among 205 proteins. These results show that BiFC-seq is a highly sensitive, rapid, and economical method for genome-wide interactome mapping.
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