Table 2.

Main approaches to the synthesis of POCs, and their advantages and limitations.

Stepwise Solid-Phase Synthesis
Conjugation via	Advantages	Disadvantages/Limitations
Bifunctional or trifunctional linker	Absence of time-consuming isolation/purification of both peptide (P) and oligonucleotide (O) fragments No excess of either P or O fragment—less solubility problems May be convenient for peptide-PNA conjugates (P-PNAs) due to protecting group compatibility	Poor compatibility of P and O chemistries: the need to design a suitable protecting group scheme. Attachment of limited number of amino acids without side-chain protection Difficulty synthesizing longer than medium-length conjugates
Post-Synthetic Conjugation
Conjugation via	Advantages	Disadvantages/Limitations
Thioether or disulfide bond	Many suitable conjugation procedures available Many reagents for functionalization of either fragment available No problem with incompatibility of the two chemistries Conjugation of peptides with any amino acid composition Conjugation of peptides of almost any length (up to proteins)	Separate multistep preparation and purification of both fragments Reaction in aqueous solvents Solubility problems with polycationic or highly hydrophobic peptides
Native ligation
Oxime, thiazolidine, or hydrazone linkage
Amide bond formation
Click chemistry
Diels-Alder reaction