Human dopamine D4 receptor allele genotyping by ultrathin agarose gel electrophoresis with To-Pro-3 complexation

Growing evidence shows the correlation between the allelic type of the dopamine D4 receptor and the human novelty-seeking personality trait. A sensitive, ultrathin agarose gel electrophoresis-based, high-throughput screening method was developed for genotyping the dopamine D4 receptor (D4DR) exon III 48 base pair repeat polymorphism. The efficiency of the method was increased by reamplification nested polymerase chain reaction (PCR) - of the 48 base pair repeat containing the PCR product with internal primers. The nested PCR fragments were analyzed by ultrathin layer agarose gel electrophoresis with an automated real-time laser-induced fluorescent detection system. Noncovalent affinity complexation was accomplished during the separation process by the addition of a very low concentration of intercalation dye, To-Pro-3 (2 nM) to the gel buffer system. This resulted in instant fluorescent labeling of the migrating PCR fragments. This method can readily facilitate genetic association studies between dopamine receptor genotypes and some human behavioral and neuropsychiatric disorders.