Multiplex Degenerate Primer Design for Targeted Whole Genome Amplification of Many Viral Genomes

Gardner, Shea N.; Jaing, Crystal J.; Elsheikh, Maher M.; Peña, José; Hysom, David A.; Borucki, Monica K.

Two multiplex reactions were set up with each containing a group of nonoverlapping primer sets (Figure 2 ). For example, multiplex "A" included primer sets A, C, E, G, and I and multiplex "B" had primer sets B, D, F, and H. By staggering the primer sets into different multiplex reactions, the amplification of overlapping primer regions created by the reverse primer from one set with the forward primer of the overlapping, adjacent primer set was eliminated. Without this strategy, these overlapping primer sets would dominate the PCR reaction due to the small size of these amplicons.