Non-radioactive direct sequencing of PCR products amplified from neuropathological specimens

We have developed a simple, rapid and relatively inexpensive protocol for direct non-isotopic cycle sequencing of DNA amplified using the polymerase chain reaction (PCR). PCR is performed on routine and archival neuropathological tissue. For sequencing, a 5'-digoxigenin end-labelled oligonucleotide primer is annealed and extended during thermal cycling, sequencing reactions are separated on a standard sequencing gel and the gel is contact-blotted to a nylon membrane. Sequenced DNA is visualized using immunological detection of digoxigenin.