PCR/SSCP detects reliably and efficiently DNA sequence variations in largescale screening projects

A simple and fast method with high reliability is necessary for the identif ication of mutations, polymorphisms and sequence variants (MPSV) within man y genes and many samples, e.g. for clarifying the genetic background of ind ividuals with multifactorial diseases. Here we review our experience with t he polymerase chain reaction/single-strand conformation polymorphism (PCR/S SCP) analysis to identify MPSV in a number of genes thought to be involved in the pathogenesis of multifactorial neurological disorders, including aut oimmune diseases like multiple sclerosis (MS) and neurodegenerative disorde rs like Parkinson's disease (PD). The method is based on the property of th e DNA that the electrophoretic mobility of single stranded nucleic acids de pends not only on their size but also on their sequence. The target sequenc es were amplified, digested into fragments ranging from 50-240 base pairs ( bp), heat-denatured and analysed on native polyacrylamide (PAA) gels of dif ferent composition. The analysis of a great number of different PCR product s demonstrates that the detection rate of MPSV depends on the fragment leng ths, the temperature during electrophoresis and the composition of the gel. In general, the detection of MPSV is neither influenced by their location within the DNA fragment nor by the type of substitution, i.e., transitions or transversions. The standard PCR/SSCP system described here provides high reliability and detection rates. It allows the efficient analysis of a. la rge number of DNA samples and many different genes.