Multiplex primer prediction software for divergent targets

Gardner, Shea N.; Hiddessen, Amy L.; Williams, Peter L.; Hara, Christine; Wagner, Mark C.; Colston, Bill W.

Primer design for amplification and detection of divergent target sequences can be challenging, and this problem will only grow as sequencing technologies improve. Some methods are limited in scalability, particularly those requiring a MSA as input. Developing a PCR multiplex is often a tedious mix-and-match process from among primers originally designed to work in singleplex. We describe the MPP algorithm based on hashing of conserved k-mer subsequences that requires no MSA and where multiplex-compatible primer sets are built de novo to avoid primer dimer and hairpin formation to the extent that can be predicted based on free energy calculations.