Direct Sequencing of PCR Products

The development of the polymerase chain reaction (PCR) has allowed the rapid isolation of DNA sequences utilizing the hybridization of two oligonucleotide primers and subsequent amplification of the intervening sequences by Taq polymerase. There are many applications of this technique. One of the most useful is the screening of large numbers of samples in the search for mutations at a defined locus, for example in clinical studies or in the analysis of cultured cell lines (1 –3 ). In the absence of PCR, this can only be achieved by isolating DNA from each individual, and making and screening a library. The use of PCR means that whereas previously it may have taken a month to examine each individual sample, it is now possible to examine many samples in a few days. In addition, by using redundant primers, it is even possible to isolate related novel genes.