A rapid screening method for a single nucleotide polymorphism (SNP) in the human MOR gene

Abstract

Aims: Genetic association studies have suggested that the single nucleotide polymorphism (SNP) at position 118 of the human mu-opioid receptor (MOR) gene could be a potential risk factor for drug treatment variability in patients. Therefore, we wanted to develop a fast and reliable detection method for this SNP which is applicable in a clinical setting.

Methods: To detect the polymorphism at position A118-->G in the human MOR gene we used the fluorescence resonance energy transfer (FRET)-PCR technique with subsequent melting curve analysis.

Results: The polymorphism at position A118-->G in the human MOR gene could be clearly discriminated with melting peak temperatures of 69.8 degrees C and 63.8 degrees C, corresponding to the wild type and mutated MOR allele, respectively. The results from FRET-PCR were validated by sequencing and restriction-fragment length polymorphism (RFLP). Screening of blood samples from 100 subjects showed an allelic distribution for the human MOR alleles of 79% (homozygous wild type), 20% (heterozygous) and 0.9% (homozygous mutated).

Conclusions: The FRET-PCR protocol for detection of the human MOR gene polymorphism at position 118 offers a rapid and reliable method which could be used for population screening of this and other genes.

Figures

Figure 1

Genotyping of the human MOR gene at position 118 with allele-specific fluorescence probes. After amplification, melting analysis was performed. Derivative melting curve plots of −dF/dT vs temperature are shown. Sample 1 is from a subject who is homozygous for the wild type MOR allele, sample 2 is from a homozygote for the mutated allele, and sample 3 is from a heterozygote. Sample 4 represents the water control.