A 2in1 cloning system enables ratiometric bimolecular fluorescence complementation (rBiFC).

摘要: Gene expression and binary interaction techniques are vital tools that shape our understanding of protein complexes. An inherent flaw, however, with most current protein-protein is the variability in levels for fusion proteins when using several individual plasmids. Here, we describe a novel recombination-based cloning strategy called 2in1 enables co-expression on cell-by-cell basis from single plasmid. We demonstrate utility through development ratiometric bimolecular fluorescence complementation assay (rBiFC), which both candidate genes simultaneously cloned into vector backbone containing an internal fluorescent marker control analysis. rBiFC significantly increases credibility results allowing comparison between different pairs. In addition to its use rBiFC, can be introduced easily other systems rely multiple gene prove feasible future synthetic biological approaches.