Giordano M, Oefner P J, Underhill P A, Cavalli Sforza L L, Tosi R, Richiardi P M
Dipartimento di Scienze Mediche, Università di Torino, Novara, Italy.
Genomics. 1999 Mar 15;56(3):247-53. doi: 10.1006/geno.1998.5715.
Genetic association analysis of candidate regions where evidence of linkage has accumulated is becoming a key issue in the study of complex diseases. A high density of markers, at least one per centimorgan, is required to improve the chances of observing linkage disequilibrium with disease alleles. A recently available single nucleotide polymorphism (SNP) map designed to cover the whole genome provides an average density of one marker per 2 cM. In the present study we show that the number of markers can be approximately doubled in a selected region, thus reaching a density suitable for association studies, by applying a completely automated technique for polymorphism detection, denaturing high-performance liquid chromatography (DHPLC). A systematic search for SNPs was performed in the region 5ptel-q13, where weak but convergent evidence for linkage with multiple sclerosis has accumulated. Screening for polymorphisms was performed on 124 sequence tagged sites (STSs) in the 3'UTR ends of expressed sequence tags totaling about 30,000 bp. Thirty SNPs in 28 STSs were found with less than 10% overlap with the markers already detected in the same region. The data confirm the validity of the approach using DHPLC on expressed gene sequences tagged by a set of standard commercially available primers.
对已积累连锁证据的候选区域进行基因关联分析正成为复杂疾病研究中的关键问题。为提高观察到与疾病等位基因的连锁不平衡的几率，需要高密度的标记，至少每厘摩一个标记。最近可用的旨在覆盖整个基因组的单核苷酸多态性（SNP）图谱提供了平均每2厘摩一个标记的密度。在本研究中，我们表明通过应用一种用于多态性检测的完全自动化技术——变性高效液相色谱法（DHPLC），在选定区域内标记数量可大约增加一倍，从而达到适合关联研究的密度。在5p端粒 - q13区域进行了SNP的系统搜索，该区域已积累了与多发性硬化症连锁的微弱但一致的证据。对总共约30,000 bp的表达序列标签3'UTR末端的124个序列标签位点（STS）进行了多态性筛选。在28个STS中发现了30个SNP，与同一区域已检测到的标记重叠率不到10%。数据证实了在一组标准商业可用引物标记的表达基因序列上使用DHPLC方法的有效性。
