Simultaneous genotyping of two SNP sites in heterozygous apoE genes by using exonuclease III, nuclease S1 and two PNA probes: (A) ε4/ε3 by PNA(S112G) and PNA(A158C) probes; (B) ε2/ε3 by PNA(S112A) and PNA(A158T) probes. The signals from codon 112 are shown by either triangles (from the C-allele(112)) or asterisks (from the T-allele(112)), whereas the signals from codon 158 are shown by either diamonds (from the C-allele(158)) or open circles (from the T-allele(158)). The signals at m/z values of 2728.1, 2744.1, 3405.4 and 3420.4 are from the PNA probes (PNA(S112A), PNA(S112G), PNA(A158C) and PNA(A158T), respectively). Reaction conditions: [dsDNA (203 bp)] = 2.5 µM, [PNA probes] = 2.5 µM, [exonuclease III] = 5.0 units/µl at pH 8.0 and 37°C for 5 min and [nuclease S1] = 3.0 units/µl at pH 4.6 and 20°C for 12 min. Mass spectra were recorded in a negative ion mode on a Bruker Daltonics® Autoflex mass spectrometer.