AcePrimer is an internet-accessed application based on CGI/Perl programming that designs PCR primers to search for deletion alleles in Caenorhabditis elegans gene knockout experiments and uses electronic PCR to search the entire genomic DNA sequence for potential false priming or multiple PCR amplification targets. Features such as the ability to target specific exons with the poison primer approach and evaluation of primers with electronic PCR provide a flexible, web-based approach to design effective primers whilst minimizing the need for empirical optimization of PCR experiments.