Integrating Two Efficient and Specific Bioorthogonal Ligation Reactions with Natural Metabolic Incorporation in One Cell for Virus Dual Labeling.

Though techniques in bioorthogonal chemistry and metabolic incorporation have been developed over the past decade, it remains difficult to integrate different bioorthogonal reactions or metabolic incorporations into one system. In this report, the protein and DNA metabolic incorporations were combined with two bioorthogonal reactions in one cell to develop a facile and universal method for virus dual labeling. Azide and vinyl groups were introduced into the proteins or genomes of viruses, respectively, through the intrinsic biosynthesis of biomolecules, which were subsequently fluorescently labeled via copper-free click chemistry or alkene-tetrazine ligation reactions during natural propagation process in host cells. Both the envelope viruses and the capsid viruses could be labeled, and the dual labeling efficiency was more than 80%. The labeled progeny virions were structurally intact and fully infectious, and their fluorescence was strong enough to track single virions.