units per sample exonuclease and 0.50 U/sample SAP, incubating for 30 min at 37° and incubating for a further 5 min at 95°. After purification, 1.5 μl of cleaned PCR product was mixed with 5.5 μl of water, 2.0 μl of SNaPshot ready Reaction Mix and 1.0 μl of mixed SNaPshot extension primers (2 pmol/μl). Twenty-five thermocycles of 96° for 10 s, 50° for 5 s, 60° for 30 s were followed by treatment of postextension products 0.50 U SAP/sample, incubation for 30 min at 37° and for 5 min at 95°. One microliter of final product was denatured with 8.5 μl Hi-Di Formamide (ABI) and 0.5 μl Liz-120 Size Standards (ABI) at 95° for 5 min. Fragments were then electrophoresed using a 3100 DNA Analyzer (ABI) and analyzed using ABI GeneScan version 3.7 software. Genotypes were scored manually, using size standards for peak verification.
