Ferraro T N, Schill J F, Ballas C, Mulholland N, Golden G T, Smith G G, Buono R J, Berrettini W H
Department of Psychiatry, Center for Neurobiology and Behavior, University of Pennsylvania, Philadelphia 19104, USA.
Psychiatr Genet. 1998 Winter;8(4):227-33. doi: 10.1097/00041444-199808040-00005.
Agarose gel electrophoresis with ethidium bromide staining (AGE/EBS) is an efficient and reliable method for analyzing microsatellite polymorphisms. We report the use of AGE/EBS for analyzing DNA microsatellite polymorphisms in a preliminary quantitative trait loci (QTL) study of seizure susceptibility in which a candidate gene strategy was used to direct initial mapping efforts. F2 intercross progeny, derived from seizure-sensitive DBA/2J (D2) and seizure-resistant C57BL/6J (B6) inbred strains of mice, were tested for their sensitivity to the seizure-inducing effect of pentylenetetrazol (PTZ), a gamma-aminobutyric acid (GABA) receptor antagonist. A semi-automated method is described, in which DNA microsatellites were amplified by polymerase chain reaction (PCR) to yield products of 100-200 base pair (bp) in length. Alleles were separated on 3-6% MetaPhor agarose gels, stained with ethidium bromide, and visualized by ultraviolet (UV) illumination. Univariate analysis of genotype and phenotype data provides evidence for a seizure-related QTL on chromosome 5, near genes coding for the GABAA receptor subunits alpha 5 and gamma 3. Interestingly, this suggestive QTL derives from the more resistant B6 strain, but it nonetheless provides impetus for the characterization of possible strain differences in these two candidate genes. Overall, these results demonstrate that AGE/EBS can be useful for rapid screening of genomic regions of special interest in QTL mapping studies.
使用溴化乙锭染色的琼脂糖凝胶电泳（AGE/EBS）是分析微卫星多态性的一种高效且可靠的方法。我们报告了在癫痫易感性的初步数量性状基因座（QTL）研究中使用AGE/EBS分析DNA微卫星多态性，该研究采用候选基因策略来指导初始定位工作。从癫痫敏感的DBA/2J（D2）和癫痫抗性的C57BL/6J（B6）近交系小鼠中获得的F2杂交后代，接受了对戊四氮（PTZ，一种γ-氨基丁酸（GABA）受体拮抗剂）诱导癫痫作用的敏感性测试。本文描述了一种半自动方法，其中通过聚合酶链反应（PCR）扩增DNA微卫星，以产生长度为100 - 200碱基对（bp）的产物。等位基因在3 - 6%的MetaPhor琼脂糖凝胶上分离，用溴化乙锭染色，并通过紫外线（UV）照射进行可视化。对基因型和表型数据的单变量分析为5号染色体上一个与癫痫相关的QTL提供了证据，该QTL靠近编码GABAA受体α5和γ3亚基的基因。有趣的是，这个提示性的QTL来自抗性更强的B6品系，但它仍然为表征这两个候选基因中可能存在的品系差异提供了动力。总体而言，这些结果表明AGE/EBS可用于在QTL定位研究中快速筛选特别感兴趣的基因组区域。
