Separate analysis of complementary strands of restriction enzyme-digested DNA. An application of restriction fragment mass mapping by matrix-assisted laser desorption/ionization mass spectrometry.
Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI/TOF-MS) of a restriction endonuclease digest determines the molecular mass of PCR-amplified DNA more easily than measurement of undigested DNA. With this method, a 664 bp region from the FAS gene could be analyzed and a two-nucleotide deletion in the L1CAM gene was detected in a restriction fragment of 105 nucleotides. Furthermore, the analysis of smaller fragments allowed separate detection of single-stranded oligonucleotides comprising individual digested fragments. This mixture analysis of restriction enzyme digests improves the resolution, sensitivity and accuracy of MALDI/TOF-MS of DNA and is thus expected to facilitate its application to genetic diagnosis.
Wada
Y
Y
Department of Molecular Medicine, Osaka Medical Center, Japan.
D004247:DNA / Q000032:analysis* / Q000235:genetics; D004262:DNA Restriction Enzymes; D006868:Hydrolysis; D009154:Mutation; D016133:Polymerase Chain Reaction; D015183:Restriction Mapping / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D004247:DNA; D004262:DNA Restriction Enzymes