Figure 2.

Representative chemical-based methods for circRNA formation. (a) An example of an EDC-based reaction [19]. The ligation for circRNA formation between 3′-amino-modified RNA and 5′-phosphorylated RNA can be completed by 1-pot EDC/HOBt chemical synthesis with a DNA splint. (b) An example of a BrCN-based reaction [20,21]. (c) An example of click chemistry using azide and alkyne for RNA circularization [22]. (d) DNA splint-based ligation [23] for pre-orientation of the 5′- and 3′-ends to be linked (red dotted circle, RNA in blue, DNA splint in gray).