PCR enhancers: Types, mechanisms, and applications in long-range PCR

Polymerase chain reaction is an important tool in molecular biology. Although the principles of the technique are relatively simple, amplifying complex or long DNA segments can be challenging. A variety of PCR additives are used to improve the performance and yield of difficult PCRs. Each PCR additive has unique properties and enhances PCR through a different mode of action. They are used to either improve PCR sensitivity, efficiency, and specificity, or mitigate the effects of PCR inhibitors. In this review, we categorise known PCR additives into four main groups. The first three groups comprise PCR additives with well-defined mechanisms, namely those that facilitate the amplification of GC-rich sequences, counteract the detrimental effects of PCR inhibitors, or alter PCR kinetics (nanomaterials). The fourth group is a loose mix of additives with unclear mechanisms of action. Then, we discuss how these additives may be used to tackle specific PCR-related challenges, particularly those associated with long-range PCR. We conclude the review with added insights into the use of PCR additives in enhancing the synthesis of complex and long DNA fragments. •The synthesis of long amplicons may benefit from some of the latest innovations in PCR.•Some additives enhance the performance of long-range PCR through specific mechanisms.•Chemical additives are underutilised in long-range PCR.•A combination of enhancers may be necessary for overcoming the challenges associated with long-range PCR.