Multiplex genotyping of the human β2-adrenergic receptor gene using solid-phase capturable dideoxynucleotides and mass spectrometry

Sobin Kim, Shundi Shi, Tomás Bonome, Michael E. Ulz, John R. Edwards, Heidi Fodstad, James J. Russo, Jingyue Ju

Research output: Contribution to journalArticle

17 Scopus citations


Previously, we established the feasibility of using solid phase capturable (SPC) dideoxynucleotides to generate single base extension (SBE) products which were detected by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for multiplex genotyping, an approach that we refer to as SPC-SBE. We report here the expanding of the SPC-SBE method as a single-tube assay to simultaneously detect 20 single nucleotide variations in a model system and 3 single nucleotide polymorphisms (SNPs) in the human β2-adrenergic receptor (β2AR) gene. Twenty primers were designed to have a sufficient mass difference between all extension products for accurate detection of nucleotide variants of the synthetic templates related to the p53 gene. These primers were extended simultaneously in a single tube with biotin-ddNTPs to generate 3′-biotinylated DNA products, which were first captured by streptavidin-coated magnetic beads and then released from the beads and analyzed with MALDI-TOF MS. This approach generates a mass spectrum free of primer peaks and their associated dimers, increasing the scope of multiplexing SNPs. We also simultaneously genotyped 3 SNPs in the β2AR gene (5′LC-Cys19Arg, Gly16Arg, and Gln27Glu) from the genomic DNA of 20 individuals. Comparison of this approach with direct sequencing and the restriction fragment length polymorphism method indicated that the SPC-SBE method is superior for detecting nucleotide variations at known SNP sites.

Original languageEnglish
Pages (from-to)251-258
Number of pages8
JournalAnalytical Biochemistry
Issue number2
StatePublished - May 15 2003
Externally publishedYes


  • Biotinylated dideoxynucleotides
  • MALDI-TOF mass spectrometry
  • Multiplex genotyping
  • SNP
  • Single base extension
  • β2-Adrenergic receptor gene

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