Core--Molecular genetics

该核心单元为每个研究项目提供关键的分子资源，包括： DNA样品制备和储存的制备和编目、基因分型、物理作图和遗传标记开发、信息支持、DNA测序以及作为候选功能变化的靶基因座处的多态性的鉴定。 1.基因组DNA（gDNA）的制备和储存。一致和可靠的DNA样本， 基因分型（分析连锁、关联和候选功能序列变化）对项目1和2至关重要。我们从诊所的每个人身上收集50 ml EDTA抗凝全血。我们从少量的白色细胞中制备gDNA，剩余的血液冷冻储存以备将来使用。 2. DNA测序战略项目1-5中的每一个都提出了生物学候选基因， 血压自主控制的遗传变异的潜在贡献者。测试这些 候选基因需要对已知标记进行连锁、关联和单倍型分析，更具体地，需要对高血压患者及其一级亲属中富集的单点和单倍型等位基因进行连锁、关联和单倍型分析。 3.基因分型。战略和方法。如上文测序部分所述，我们使用两层策略来最大限度地提高效率。(1)第一层涉及候选基因座的直接基于PCR的重测序，以提高密度和多态性选择的适当性，用于更大规模的基因分型。 (2)在策略的第二层中，选择衍生的标记组合并用于允许有效区分单倍型。随后在信息学核心B（信息学和统计遗传学）中分析获得的基因型数据的统计学显著性和群体参数，包括可能准确性的额外的基于群体的对照。 4.等位基因特异性表达分析。战略为了检测本程序中定义的等位基因变体对表达水平的影响，该核心将提供基于基因分型样品和异源启动子融合的等位基因检测的表达分析。

This core unit provides crucial molecular resources for each of the research projects, including: preparation and cataloging of DNA sample preparation and storage, genotyping, physical mapping and genetic marker development, informatic support, DNA sequencing, and identification of polymorphisms at targeted loci as candidate functional changes. 1. Preparation and storage of genomic DNA (gDNA). Consistent and reliable DNA samples for genotyping (in analyses of linkage, association, and candidate functional sequence changes) are essential to Projects 1 and 2. We collect 50 ml of EDTA-anticoagulated whole blood from each individual in the clinic. We prepare gDNA from the white cells in small amounts and the remaining blood is stored frozen for future use. 2. DNA Sequencing. Strategy. Each of Projects 1-5 has suggested biological candidate genes as potential contributors to genetic variation in autonomic control of blood pressure. Testing these candidate genes requires linkage, association, and haplotype analysis of known markers and more particularly of single point and haplotype alleles enriched in hypertensive subjects and their first-degree relatives. 3. Genotyping. Strategy and Methods. As described under Sequencing above, we use a two-tiered strategy to maximize efficiency. (1) The first tier involves direct PCR-based resequencing of candidate loci to improve the density and appropriateness of polymorphism selection for larger-scale genotyping. (2) In the second tier of the strategy, derived marker combinations are selected and used to allow efficient discrimination of haplotypes. Obtained genotype data is subsequently analyzed for statistical significance and population parameters in the informatics core, Core B (Informatics and statistical genetics), including additional population-based controls for likely accuracy. 4. Allele-specific expression analysis. Strategy. To detect the effects of allelic variants define in this program on expression level, this core will provide expression analysis based on allelic detection from genotyped samples and heterologous promoter fusions.