Genotyping single-nucleotide polymorphisms by matrix-assisted laser desorption/ionization time-of-flight-based mini-sequencing.
Currently, there is a critical need to develop high-throughput, low-cost, and accurate methods for genotyping of single-nucleotide polymorphisms (SNPs). The matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometrically based technique represents a new promising approach to SNP analysis. We have developed a new MALDI-TOF-based mini-sequencing assay, termed "VSET," for genotyping of SNPs. In this assay, specific fragments of genomic DNA containing the SNP site(s) are first amplified, followed by mini-sequencing in the presence of three ddNTPs and the fourth nucleotide in the deoxy form. In this way, the primer is extended by only one base from one allele, whereas it is typically extended by two bases from another allele. The products are then analyzed using MALDI-TOF mass spectrometry. The genotype of the SNP site is identified based on the number of nucleotides added.
Sun
Xiyuan
X
Department of Chemistry, Cleveland State University, OH, USA.
Guo
Baochuan
B
D004252:DNA Mutational Analysis / Q000379:methods; D005838:Genotype; D006801:Humans; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*