The emergence of bioorthogonal reactions has greatly advanced research in the fields of biology and medicine. They are not only valuable for labeling, tracking, and understanding biomolecules within living organisms, but also important for constructing advanced bioengineering and drug delivery systems. As the systems studied are increasingly complex, the simultaneous use of multiple bioorthogonal reactions is equally desirable. In this review, we take a look at the different bioorthogonal reactions that have recently been developed, the methods of cellular incorporation and the strategies to create orthogonality within the bioorthogonal landscape.