Single-Strand Conformation Polymorphism (SSCP) Can Be Explained by Semistable Conformation Dynamics of Single-Stranded DNA

Abstract

The cause of separation in gel electrophoresis between highly homologous ss (single-stranded) DNAs as observed in SSCP (single-strand conformation polymorphism) was pursued. Advancing our previous explanation [J. Biochem. 99, 663-671 (1986)], the mobility difference of ssDNAs was correlated to differences in their dynamic conformation (not to differences in their most stable structure), focusing on point-substituted sites. The contribution of semistable conformation dynamics was considered to be critical. Putative factors which may influence mobility (i.e., length of ssDNAs, location of substituted sites, and types of substitutions) were experimentally examined and critically discussed. Understanding of these phenomena should yield improvements in various techniques, such as SSCP, and in evaluation of the solution structures of DNA.

A comparison of fluorescent SSCP and denaturing HPLC for high throughput mutation scanning

“…Attempts to predict changes in SSCP mobility by modelling single stranded DNA conformations using the structure prediction program Mfold (http:// www.ibc.wustl.edu/~zuker/) did not give consistent results (data not shown). While it is possible that refinements in modelling the structures of single stranded DNA will lead to better predic-tions of SSCP mobility [Nakabayashi and Nishigaki, 1996], the idiosyncratic nature of SSCP remains its greatest weakness as a diagnostic tool [Liu et al, 1999].…”

Dynamics of physiologically relevant noncanonical DNA structures: an overview from experimental and theoretical studies

“…A slight alteration in the sequence (few point mutations) causes mutated DNA strands to attain a different predominant semi-stable secondary structural form or a structure similar to the wildtype ssDNA with subtle variations in the conformation. Through a set of elegant experiments, Nishigaki et al and others showed that this conformational polymorphism among different ssDNA sequences results in significantly different electrophoretic migration and differential sensitivities to enzymatic cleavage [48][49][50][51]. Currently, over 2000 genes are known to carry at least one pathogenic mutation, and the number is steadily rising [52].…”

Single-stranded DNA (ssDNA) production in DNA aptamer generation

“…The smeary appearance of ssDNA on the native PAGE due to different conformations of three-dimensional structures makes the selective excision of ssDNA tedious. [85][86][87] However, the problem of incomplete digestion can be tackled during manufacturing by ensuring that the primer is phosphorylated to the highest degree possible. Upon the completion of synthesis, the blocking group which temporarily protects the phosphate group throughout the oligonucleotide synthesis must be completely removed.…”