An MspI polymorphism in the human acid sphingomyelinase gene (SMPD1).

Protocol: To detect the 506 polymorphism, a 567 bp SMPD1 genomic fragment is amplified using sense (5'-AGTAGTCGACATGGGCAGGATGTGTGG-3') and antisense (5'-AGTAGTGTCGACTTGCCTGGTTGAACC AC AGC-3') primers. Dot-blot hybridization is performed using allele-specific oligonucleotides for 506-Arg (5'-ACTACTCCAGGAGCTCT-3') and for 506-Gly (5'-ACTACTCCGGGAGCTCT-3'), which are hybridized at 42°C and washed at 51 or 53°C, respectively. This polymorphism also can be detected by restriction enzyme analysis since the 506-Gly polymorphism creates a new Mspl restriction site. When the 567 bp PCR-amplified genomic fragment from a 506-Arg allele is digested with Mspl, two fragments of 395 and 159 bp are detected on a 1% agarose gel. In contrast, amplification of the 506-Gly allele results in the constant fragment of 395 bp, but the 159 bp fragment is digested into two fragments of 95 and 64 bp.