Efficient SNP analysis enabled by joint application of the muTGGE and heteroduplex methods.
Gene science-based diagnoses have become an increasingly realistic option as the state of knowledge has improved regarding the genetic basis of disease. To facilitate the creation of this potential diagnostic tool, researchers have made large-scale detection of point mutations a key issue. Here, we propose an inexpensive and convenient method with a high performance level for this purpose: micro temperature gradient gel electrophoresis (muTGGE)-empowered heteroduplex analysis (muTG-HD). First, muTGGE was shown to separate double-stranded DNA containing single nucleotide polymorphism (SNP) with sufficiently high resolution when used in the mode of perpendicular TGGE. Using human c-Ki-ras and rat p53 DNA, point mutations could be unequivocally detected by muTG-HD when parallel TGGE was employed. The mutation type (such as G/C to A/T), the position of the point mutation (centre or not) and the DNA size (around 100 or 200 bp) were examined and found to be detectable. Thus, muTG-HD could detect point mutations efficiently at a much lower cost by having multiple lanes per gel.
Salimullah
Md
M
Department of Functional Materials, Saitama University, 255 Shimo-Okubo, Sakura-ku, Saitama-shi, Saitama 338-8570, Japan.
Hamano
Keiichi
K
Tachibana
Masayoshi
M
Inoue
Kinji
K
Nishigaki
Koichi
K
D000818:Animals; D004252:DNA Mutational Analysis; D004591:Electrophoresis, Polyacrylamide Gel; D016158:Genes, p53 / Q000235:genetics; D011905:Genes, ras / Q000235:genetics; D020180:Heteroduplex Analysis / Q000295:instrumentation / Q000379:methods*; D006801:Humans; D016297:Mutagenesis, Site-Directed; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*; D051381:Rats