Establishment of genetic diagnosis protocol for color blindness using oral mucosa scratch sample

色盲是一种X连锁遗传病，影响大约5%-10%的男性。据估计，女性携带者约占日本人口的10%。在没有家族谱系信息的情况下，基因分析是识别女性携带者的唯一方法。本研究的目的是建立一种可靠的方案来识别女性携带者。DNA样本取自志愿者口腔黏膜拭子。采用改良的组织基因组DNA提取方法提取基因组DNA。β-肌动蛋白基因的阳性对照聚合酶链式反应显示，大肠粘膜拭子标本扩增成功。设计视蛋白基因的引物，用于聚合酶链式反应分析。虽然获得了几个扩增产物，但DNA测序结果表明这些产物都是非特异性扩增。这一结果表明，要想进行有效的扩增，还需要对引物设计进行进一步的改进。从口腔黏膜拭子样本中成功地扩增了其他几个基因，因此我们的样本收集和DNA提取方法被认为是足够和可靠的用于诊断的。

Color blind is an X-linked genetical disease that affects approximately 5-10 % males. Female carrier is estimated to count approximately 10 % of the Japanese population. Gene analysis is the only way to identify a female carrier when family pedigree information is not available. The purpose of the present study is to establish a reliable protocol to identify a female carrier. DNA sample was obtained from buccal mucosa swab of volunteers. Genomic DNA was extracted using modified protocol for tissue genomic DNA extraction. Positive control PCR for beta-actin gene revealed successful amplification from buocal mucosa swab samples. Primers for opsin gene was designed and used for PCR analysis. Although several PCR products were obtained, DNA sequencing results showed that these products were non-specific amplification. This result indicate sthat further refinement of primer design is require for efficient amplification. Several other genes were successfully amplified from buccal mucosa swab samples, thus our sample collection and DNA extraction protocol was thought to be sufficient and reliable for diagnostic use.