COMPREHENSIVE DNA DIAGNOSTIC SYSTEM FOR SINGILE GENE DISORDERS

本研究的目的是建立多种单基因疾病的综合DNA诊断体系。为此，我们使用了一种新的DNA试纸法来检测先前鉴定的致病突变。我们还使用变性高效液相色谱(DHPLC)扫描未知突变，并使用自动毛细管DNA测序仪确认突变。DNA试纸法最初是由我们实验室开发的，它利用竞争性等位基因特异的短寡核苷酸杂交和免疫层析试纸相结合。它被用于检测糖原储存疾病Ia和Ib、非酮症高血糖、乙醛脱氢酶2缺乏、Tay-Sachs病、Gaucher病、细胞周期纤维化、因BRCA1或BRCA2突变引起的家族性乳腺癌、因子V Leiden突变、CYP2C19和线粒体A1555G突变等已知突变。我们还从临床诊断的角度评估了DHPLC和测序分析的可行性。这三种方法的结合将通过有效地识别突变来促进DNA诊断。

The purpose of the current study was to establish comprehensive DNA diagnostic system for various single gene disorders. To this end, we used a novel DNA test strip method to detect previously idertified pathogenic mutations. We also used denaturing high-performance liquid chromatography (DHPLC) to scan for unknown mutations and an automatic capillary DNA sequencer to confirm mutations. The DNA test strip method, originally developed in our laboratory, utilizes competitive allele-specific short oligonucleotide hybridization coupled with immunochromatographic strip. It was used for the detection of known mutations in glycogen storage disease type Ia and Ib, non-ketotic hyperglycinemia, aldehyde dehydrogenase 2 deficiency, Tay-Sachs disease, Gaucher disease, cyctic fibrosis, familial breast cancer due to BRCA1 or BRCA2 mutations, Factor V Leiden mutation, CYP2C19 and mitochondrial A1555G mutation. We also evaluated the feasibility of DHPLC followed by sequencing analysis from clinical diagnostic point of view. The combination of the three methods would facilitate DNA diagnosis by efficiently identifying mutations.