MALDI mass spectrometry analysis of single nucleotide polymorphisms by photocleavage and charge-tagging.
High-throughput procedures are an important requirement for future large-scale genetic studies such as genotyping of single nucleotide polymorphisms (SNPs). Matrix-assisted laser desorption/ ionisation mass spectrometry (MALDI-MS) has revolutionised the analysis of biomolecules and, in particular, provides a very attractive solution for the rapid typing of DNA. The analysis of DNA by MALDI can be significantly facilitated by a procedure termed 'charge-tagging'. We show here a novel approach for the generation of charge-tagged DNA using a photocleavable linker and its implementation in a molecular biological procedure for SNP genotyping consisting of PCR, primer extension, photocleavage and a chemical reaction prior to MALDI target preparation and analysis. The reaction sequence is amenable to liquid handling automation and requires no stringent purification procedures. We demonstrate this new method on SNPs in two genes involved in complex traits.
Sauer
Sascha
S
Max-Planck-Institute for Molecular Genetics, Abteilung Lehrach, Ihnestrasse 73, 14195 Berlin-Dahlem, Germany. sauer@molgen.mpg.de
Lehrach
Hans
H
Reinhardt
Richard
R
D017931:DNA Primers; D005838:Genotype; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D020641:Polymorphism, Single Nucleotide; D017422:Sequence Analysis, DNA / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013873:Thionucleotides / Q000737:chemistry; D014466:Ultraviolet Rays
D017931:DNA Primers; D009838:Oligodeoxyribonucleotides; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D013873:Thionucleotides