Detection of differences in higher order structure between highly homologous single-stranded DNAs by low-temperature denaturant gradient gel electrophoresis.

Denaturant gradient gel electrophoresis performed at low temperature was shown to be able to detect the mobility change corresponding to the denaturation of the secondary structure of single-stranded(ss)DNA. Mobility transitions observed were determined to correspond to the melting of local structures, since both denaturants and temperature were verified to have similar effects on the mobility transition of single-stranded DNAs as on that of double-stranded DNAs. In this study it was found that point mutations can effectively change the secondary structures of ssDNA and RNA, and that the method adopted here is very sensitive to such alterations. The validity of the method was supported by a computer analysis of the secondary structure of DNA.