NONRADIOACTIVE 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 po lymerase chain reaction (PCR). PCR is performed on routine and archiva l neuropathological tissue. For sequencing, a 5'-digoxigenin end-label led oligonucleotide primer is annealed and extended during thermal cyc ling, sequencing reactions are separated on a standard sequencing gel and the gel is contact-blotted to a nylon membrane. Sequenced DNA is v isualized using immunological detection of digoxigenin.