Boronate esters, formed by the reaction of an oligonucleotide bearing a 5′‐boronic acid moiety with the 3′‐terminal cis‐diol of another oligonucleotide, support the assembly of functional nucleic acid architectures. Reversible formation of boronate esters occurs in templated fashion and has been shown to restore the activity of split DNA and RNA enzymes as well as a split fluorescent light‐up aptamer. Apart from their suitability for the design and application of split nucleic acid enzymes and aptamers in the field of biosensing, boronate esters may have played an important role in early life as surrogates of the natural phosphodiester bond. Their formation is reversible and thus fulfills an important requirement for biological self‐assembly. Here we discuss the general concept of stimuli‐dependent boronate formation and its application in biomolecules with implications for future research.