Single strand conformation polymorphism (SSCP) analysis remains one of the most widely used techniques to screen for unknown genetic variants prior to more expensive DNA sequencing. Although SSCP is cost effective and relatively simple to perform, the sensibility and reproducibility of this method rely critically on the experimental conditions during the electrophoretic run.
One of the better devices to achieve optimal SSCP results is the GenePhor Electrophoresis Unit from GE Healthcare (formerly Amersham Biosciences). The GenePhor system consists of a horizontal gel unit with an integrated Peltier cooling system that allows for very accurate temperature control over the entire gel surface. Additionally, the device works with pre-cast polyacrylamide gels, as well as, semi-dry buffer strips, totally eliminating the need for time-wasting liquid handling. The efficient cooling system combined with the use of pre-cast gels (available in a variety of polyacrylamide concentrations) ensures a rapid optimization of the experimental conditions and a high run-to-run reproducibility. Unfortunately, precast gels are not cheap; 24 PCR samples may be run simultaneously at a cost of about $15 per gel.
In the last two years, I have used the GenePhor Electrophoresis Unit to help me screen for mutations in the entire coding region (38 exons) of the ATP-binding cassette C9 (ABCC9) gene. I empirically tested different experimental conditions and found that only two different temperatures (5° and 12°) were required to obtain clean band patterns when analyzing PCR products up to 350 base pairs. Under these conditions, I have been able to identify a total of 12 different known and unknown sequence variations in this gene.
Overall, I would say that the GenePhor Electrophoresis Unit is an excellent, easy-to-use platform that allows quick and reproducible results when scanning for unknown sequence variants. Overall, I would suggest that one should consider accepting the cost of precast gels, rather than compromise on mutation detection efficiency.
Enzo Emanuele
MD
Interdepartmental Center for Research in Molecular Medicine
University of Pavia, Italy