Analysis of DNA fragments from conventional and microfabricated PCR devices using delayed extraction MALDI-TOF mass spectrometry.
Applications of polymerase chain reaction (PCR) product analysis using rapid affinity capture followed by delayed extraction (DE) MALDI-TOFMS is presented. Such applications include multiplex short tandem repeat (STR) typing, which is demonstrated for STR systems from conventional and microchip-based thermal cycling instruments. Using the combination of the microfabricated PCR instrument and DE-MALDI-TOFMS, a complete genotyping assay can be performed in under 50 min with a resultant molecular weight accuracy approaching or exceeding 100 ppm through external calibration. The observed resolution and mass accuracy for a 69-base PCR product enables identification of single base substitutions by direct molecular weight determination.
Ross
P L
PL
DNA Technology Development Branch, Center for Medical and Molecular Genetics, Armed Forces Institute of Pathology, Rockville, Maryland 20850, USA. philip_ross@pbio.com
Davis
P A
PA
Belgrader
P
P
D002138:Calibration; D004247:DNA / Q000032:analysis* / Q000737:chemistry; D053938:DNA Fragmentation; D005838:Genotype; D006801:Humans; D008970:Molecular Weight; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic; D012091:Repetitive Sequences, Nucleic Acid; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
D004247:DNA