In this article, two engineering-strategies were carried out to enhance the processivity of the DNA polymerase used in recombinase polymerase amplification (RPA). We demonstrate that covalent linkage of a non-specific, double-stranded DNA binding protein, Sso7d, to the large fragment of Staphylococcus aureus Pol I (Sau) caused a moderate enhancement of processivity and a significant improvement in the salt tolerance of Sau. Meanwhile, we provide evidence suggesting that insertion of the thioredoxin-binding domain from bacteriophage T7 DNA polymerase into the analogous position of the large fragment of Sau dramatically enhanced the processivity and mildly increased the salt tolerance of Sau when a host DNA binding protein, thioredoxin, was annexed. Both of these two strategies did not improve the amplifying performance of Sau in RPA, indicating that optimum processivity is crucial for amplifying efficiency.
Keywords: Processivity; Recombinase polymerase amplification; Salt tolerance; Sso7d; Thioredoxin-binding domain.