Abstract
Single-strand conformational polymorphism (SSCP) analysis is a technique used to screen for the presence of sequence variations in short DNA fragments. This technique relies on the ability of single-stranded DNA molecules to fold into unique secondary structures, the conformations for which are based on their primary nucleotide sequence. Changes in the nucleotide sequence, owing to a polymorphism or a mutation, are expected to alter the secondary structure of the molecule resulting in a shift in mobility through a nondenaturing polyacrylamide gel. It is this aberrant migration pattern that indicates the presence of a DNA sequence alteration. SSCP analysis is capable of detecting single nucleotide differences and, since its first use by Orita et al. (1,2) this technique has been extremely successful and widely used to detect disease-causing mutations (3–8). Although SSCP analysis has experienced wide spread use, its sensitivity is variable and has been reported to range from 35–100 % (9). Overall, however, the sensitivity of SSCP analysis reported in the literature usually ranges from 75–98 % with the most critical parameter being the size of the DNA fragment being evaluated. There are numerous reports stating that the sensitivity of SSCP analysis decreases as the size of the polymerase chain reaction (PCR) products become larger than 200 base pairs (10,11). Other parameters such as electrophoresis temperature, buffer concentration, gel concentration, cross-linker concentration, and the addition of compounds to the gel matrix, have also been altered to optimize SSCP sensitivity.
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Scoggan, K.A., Bulman, D.E. (2003). Single-Strand Conformational Polymorphism Analysis (SSCP) and Sequencing for Ion Channel Gene Mutations. In: Potter, N.T. (eds) Neurogenetics. Methods in Molecular Biology™, vol 217. Springer, Totowa, NJ. https://doi.org/10.1385/1-59259-330-5:143
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DOI: https://doi.org/10.1385/1-59259-330-5:143
Publisher Name: Springer, Totowa, NJ
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