Why did my PCR reaction stop working?

If your PCR reaction used to work and then stopped, the primers or template may have degraded. The primers can last up to 1 year if stored lyophilized and aliquoted at -20°C. Freezing and thawing cause the primers degradation. Primers or template may also degrade due to contamination. Millipore filtration systems, which require the filter to be changed periodically, can supply water of questionable quality at the end of a cycle. If you are using a filter system, check to see if your filter was changed soon after resuspending your oligos. If this is the case, you may need to consider changing the filters more often.