Teupser Daniel, Heino Norman, Wilfert Wolfgang, Thiery Joachim
Institute of Laboratory Medicine, Clinical Chemistry and Molecular Diagnostics, University Hospital Leipzig, Liebigstr. 27, 04103 Leipzig, Germany.
J Neurosci Methods. 2002 Mar 30;115(1):93-6. doi: 10.1016/s0165-0270(02)00008-0.
The common single nucleotide polymorphism at codon 129 of the prion protein gene is a key determinant of the genetic susceptibility to Creutzfeldt-Jakob disease (CJD). Recently, a molecular classification of sporadic CJD based on the M129V genotype in conjunction with other determinants was proposed. In the present study, we describe the development and evaluation of a rapid fluorescent-based assay to detect this polymorphism using the LightCycler system. The two polymorphic alleles could be clearly distinguished by their melting points at 52.1 and 60.4 degrees C, representing the 129V and 129M alleles, respectively. These results were confirmed by DNA sequencing. We evaluated our test in 400 patient samples and found no deviations from the expected melting patterns. The calculated allele frequency for the M-allele was 0.66. Thus, we have established a rapid, reliable fluorescent assay for high-throughput detection of the prion protein M129V polymorphism.
朊蛋白基因第129密码子处常见的单核苷酸多态性是克雅氏病（CJD）遗传易感性的关键决定因素。最近，有人提出了一种基于M129V基因型并结合其他决定因素的散发性CJD分子分类方法。在本研究中，我们描述了一种使用LightCycler系统基于荧光的快速检测方法的开发与评估，用于检测这种多态性。通过52.1℃和60.4℃的熔点可以清楚地区分两个多态性等位基因，分别代表129V和129M等位基因。这些结果通过DNA测序得到了证实。我们在400份患者样本中对我们的检测方法进行了评估，未发现与预期熔解模式有偏差。计算得出M等位基因的频率为0.66。因此，我们建立了一种快速、可靠的荧光检测方法，用于高通量检测朊蛋白M129V多态性。
