Tahira Tomoko, Kukita Yoji, Higasa Koichiro, Okazaki Yuko, Yoshinaga Aki, Hayashi Kenshi
Division of Genome Analysis, Research Center for Genetic Information, Medical Institute of Bioregulation, Kyushu University, Fukuoka, Japan.
Methods Mol Biol. 2009;578:193-207. doi: 10.1007/978-1-60327-411-1_12.
The single strand conformation polymorphism (SSCP) method is a sensitive technique used to detect subtle sequence differences in PCR-amplified DNA fragments as separated peaks in electrophoretic analysis. In this chapter, we focus on SSCP analysis for quantifying polymorphic alleles rather than scanning for mutations. Short fragments carrying single nucleotide polymorphisms are amplified from individual and pooled DNA samples, then the products are labeled with fluorescent dyes and analyzed by automated capillary electrophoresis under nondenaturing conditions. Dedicated software, QSNPlite, interprets trace data of the electrophoresis to identify alleles of individuals and quantify these alleles in the pool. The software can also incorporate sequencing data to assign alleles at the nucleotide level. The procedures described here are being used in association studies that compare allele frequencies between cases and controls to identify genes responsible for common diseases.
单链构象多态性（SSCP）方法是一种灵敏的技术，用于检测PCR扩增的DNA片段中的细微序列差异，这些差异在电泳分析中表现为分离的峰。在本章中，我们重点关注用于定量多态性等位基因的SSCP分析，而非扫描突变。从个体和混合DNA样本中扩增携带单核苷酸多态性的短片段，然后用荧光染料标记产物，并在非变性条件下通过自动毛细管电泳进行分析。专用软件QSNPlite可解释电泳的痕量数据，以识别个体的等位基因并在混合样本中对这些等位基因进行定量。该软件还可以整合测序数据，在核苷酸水平上确定等位基因。这里描述的程序正用于病例与对照之间比较等位基因频率以识别导致常见疾病的基因的关联研究中。
