Cycle Sequencing of Polymerase Chain Reaction-Amplified Genomic DNA with Dye-Labeled Universal Primers

Cycle sequencing of polymerase chain reaction (PCR) products has become the method of choice for sequencing known genomic DNA fragments. When fluorescent DNA sequencers are used, this requires the incorporation of a fluorescent dye into the cycle sequencing product either by using labeled sequencing primers or labeled terminators. The dye terminator approach is convenient because the same, unlabeled primers can be used for PCR amplification and cycle sequencing, and is cost efficient, particularly when a PCR product has to be sequenced a few times only. However, this method is not available for all DNA sequencers. Synthesizing specific dye-labeled primers, on the other hand, is expensive, and it is generally not affordable to label all PCR primers that at some stage one might like to use for sequencing.