Designing PCR Primers to Amplify Specific Members or Subgroups of Multigene Families

Multigene families throw a particular twist into the problem of polymerase chain reaction (PCR) primer design. One frequently needs primers that will not only amplify the sequence of interest, but at the same time fail to amplify the set of very similar sequences that comprises the remaining members of the family. In a “normal” PCR experiment, one must worry about “random” priming on unrelated sequences. With multigene families, one must attempt to avoid nonrandom priming on related sequences by exploiting differences that distinguish the sequence of interest. Most “primer design” programs do not help with this problem. Here we discuss what is involved in designing subfamily-specific primers.