Haddad L, Day L B, Attwood J, Povey S, Humphries S E, Day I N
Department of Medicine, Rayne Institute, University College London Medical School.
Ann Hum Genet. 1997 Nov;61(Pt 6):497-506. doi: 10.1046/j.1469-1809.1997.6160497.x.
Co-segregation studies based on a selection of intragenic restriction fragment length polymorphisms of the low density lipoprotein receptor (LDLR) gene have been used extensively both for research and diagnostic studies of familial hypercholesterolaemia (FH) families, because direct mutation screening remains complex. Here we describe the development and application of a more efficient approach to co-segregation studies based on highly informative dinucleotide and tetranucleotide repeats flanking the LDLR gene. A series of microsatellites (D19S391, D19S394, D19S221 and D19S179) were selected for study on the basis of linkage analysis in the CEPH families using intragenic polymorphisms for a TA repeat (exon 18) in the LDLR gene, and earlier data for a Pvu II polymorphism (intron 15). A physical map of the region of chromosome 19 also contributed to this selection. One marker in particular, D19S394, sited 150 kilobases telomeric to the gene, was extremely useful, displaying 90% heterozygosity, robust PCR of tetranucleotide repeats without stutter bands, and no recombination with the LDLR gene (theta = 0, LOD 68). Use of this marker in the families of twenty-three FH probands from Hampshire demonstrated co-segregation of the hyperlipidaemia phenotype with the LDLR gene region, except in one family with defective apolipoprotein B-100, and a family turning out to display familial combined hyperlipidaemia. This approach should facilitate the search for any families where FH does not co-segregate with the LDLR gene, and will enhance the repertoire of molecular diagnostic tools available for FH.
由于直接突变筛查仍然很复杂，基于低密度脂蛋白受体（LDLR）基因的一些基因内限制性片段长度多态性进行的共分离研究已广泛用于家族性高胆固醇血症（FH）家族的研究和诊断。在此，我们描述了一种基于LDLR基因侧翼高信息含量二核苷酸和四核苷酸重复序列的更有效共分离研究方法的开发和应用。基于使用LDLR基因中TA重复序列（外显子18）的基因内多态性以及先前关于Pvu II多态性（内含子15）的数据，在CEPH家族中进行连锁分析，选择了一系列微卫星（D19S391、D19S394、D19S221和D19S179）进行研究。19号染色体区域的物理图谱也有助于这一选择。特别是位于该基因端粒150千碱基处的一个标记D19S394非常有用，其杂合度为90%，四核苷酸重复序列的PCR扩增稳定，无拖带，且与LDLR基因无重组（θ = 0，LOD 68）。在来自汉普郡的23名FH先证者的家族中使用该标记进行研究，结果表明除了一个载脂蛋白B - 100缺陷的家族和一个显示家族性混合性高脂血症的家族外，高脂血症表型与LDLR基因区域共分离。这种方法应有助于寻找FH与LDLR基因不共分离的任何家族，并将增加可用于FH的分子诊断工具的种类。
