Detection of single-nucleotide mutations including substitutions and deletions by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

✍ Scribed by Y. Wada; M. Yamamoto

• Publisher: John Wiley and Sons
• Year: 1997
• Tongue: English
• Weight: 119 KB
• Volume: 11
• Category: Article
• ISSN: 0951-4198
• DOI: 10.1002/(sici)1097-0231(19971015)11:15<1657::aid-rcm83>3.0.co;2-1

✦ Synopsis

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) was applied to the detection of mutations involving single nucleotides. For detection of a single-nucleotide deletion, a normally 44 bp region of the L1CAM gene was amplified in a 50 microL solution, and measurement was carried out on the DNA sample after phenol extraction and ethanol precipitation. A molecular mass decrease of 300 Da corresponding to a single nucleotide was identified in the amplified product of patient DNA. For detection of a substitution, an amplified product from a 50 bp region of the human beta-globin gene was cleaved with restriction endonucleases HaeIII and Bsp12861. Measurement of a mixture of digested fragments, or restriction fragment mass mapping, clearly identified a heterozygous G/C mutation in the molecular ion signals for both sense and antisense single-stranded DNAs. The results indicate that MALTI-TOFMS is feasible for genetic diagnosis of point mutations.