Research on unknown pigmentary disorders caused by mutaiton of genes for melanogenesis.

等位基因特异性扩增（ASA）是一种简单的、无放射性的技术，用于检测产生遗传疾病的已知点突变。虽然这项技术是基于用等位基因特异性引物聚合酶链反应（PCR）对目标等位基因进行特异性扩增，但扩增的特异性可能取决于不同的PCR条件。为了避免非特异性扩增导致ASA的假阳性结果，我们修改了正常和突变等位基因特异性引物，使它们在倒数第二个3'位置上有一个恒定的碱基错配。通过使用酪氨酸酶阴性眼皮肤白化病（OCA）患者的基因组dna作为模板，我们证实了我们的修饰可以抑制这种不利的扩增。然后我们分析了酪氨酸酶阴性OCA的新患者。并根据临床检查结果和使用改良等位基因特异性引物的ASA毛球测试结果进行诊断。结果表明，日本患者的酪氨酸酶基因存在3个以上的病理突变等位基因。

Allele-specific amplification (ASA) is a simple and non-radioactive technique for detecting known point mutations that produce genetic diseases. Although this technique is based on the specific amplification of the target allele by a polymerase chain reaction (PCR) with allele-specific primers, the specificity of the amplification may depend on various PCR conditions. To avoid non-specific amplification which leads to false-positive results in ASA,we modified both the normal and mutant allele-specific primers so that they would have one constant base mismatch, located at the penultimate 3' position. We confirmed that our modification could inhibit such unfavorable amplification by using as templates genomic DNAs of patients affected with tyrosinase-negative oculocutaneous albinism (OCA). We then analyzed new patients affected with tyrosinase-negative OCA.and based the diagnosis on both the results of a clinical examination and those of a hair bulb test using ASA with the modified allele-specific primers. The results indicated that more than 3 alleles of the tyrosinase gene with a pathological mutation existed in Japanese patients.