Analysis of single-strand DNA conformation polymorphism by capillary electrophoresis.
Analysis of single-strand conformation polymorphism (SSCP) by capillary electrophoresis (CE) was developed. The conformational change of single-strand DNA is caused by a mutation in a DNA fragment. The change is detected as mobility shift in CE. The effects of acrylamide gel concentration, running temperature and fragment size amplified by the polymerase chain reaction (PCR) were studied to develop the separation of SSCP. The model DNA used was the divE 42 gene carrying wild- and mutant-type (G-->A point mutation at the 141 site). The results show that two single-strand DNA fragments that differ in one nucleotide can be separated by CE within minutes. This method was also applied to the separation of SSCP for N-ras gene including four kinds of mutations. All mutations tested in this study could be distinguished. CE is well suited for clinical analysis of SSCP because it is rapid and reproducible, allows on-line detection and is easy.
Arakawa
H
H
School of Pharmaceutical Sciences, Showa University, Tokyo, Japan.
Nakashiro
S
S
Maeda
M
M
Tsuji
A
A
D001483:Base Sequence; D004269:DNA, Bacterial / Q000032:analysis* / Q000737:chemistry; D019075:Electrophoresis, Capillary; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics; D005798:Genes, Bacterial; D011905:Genes, ras; D008969:Molecular Sequence Data; D009154:Mutation; D016133:Polymerase Chain Reaction; D018807:Polymorphism, Single-Stranded Conformational; D013696:Temperature
D004269:DNA, Bacterial