GENETIC AND PHYSICAL ANALYSIS OF RESTRICTION LANDMARKS

小鼠和人类基因组中的功能基因与CpG相关 岛屿。目前的估计表明，可能有多达4.5倍的 人类基因组中有10/4的岛屿，而老鼠基因组中的岛屿略少。 这些CpG岛与两个普遍表达的 (管家)基因以及具有组织特异性表达和 时间规章制度。限制性标志性基因组扫描方法 (RLGS)通过以下方式将基因组中罕见的限制位点识别为标志物 酶消化后的同位素末端标记。这些末端标记的 片段在琼脂糖凝胶中分离，然后用 第二种酶。分离的双酶切片段在凝胶中被消化 使用第三种限制性内切酶，结果产物被分离 在丙烯酰胺的最终维度中。这些地标的遗传变异 已经在近亲交配的实验室小鼠品系和 常见的实验室菌株和不同的小鼠物种。变化中的 可以使用任一标准杂交在小鼠基因组中定位标志 或重组近交系分析。遗传变异可以用来 在其他实验环境中，遵循父母特定的等位基因。为 例如，CpG岛的甲基化与 导致表观遗传的基因组印记现象 针对不同基因的亲本特异性等位基因的修改导致 单调的表情。父母特有的影响也很重要 与肿瘤发生相关的基因组不稳定性的分析。损失- 杂合性缺失(LOH)与多种肿瘤相关。一个 扫描基因组以发现这类事件的简单方法是 基因组分析的重要实验应用。在类似的情况下 时尚，这些地标的遗传变异可以作为标记 对复杂或多位点性状的遗传分析。在这种情况下，老鼠 从分离的表现出特定表型的杂交组合中 减少基因鉴定实验工作量的分析 受影响的后代所共有的差异。最后，它是 可以使用多种酶组合的RLGS分析进行基因组分析 用于识别候选基因的区域，如同源菌株 染色体区域。这是对先前提案的修订应用 对小鼠基因组中的RLGS基因座进行遗传定位。我们的主要目标是 这项建议是为了完成进口RLGS的初步阶段 方法从日本传入我们的实验室，并将其确立为 分析与CpG相关的地标的实验方法 基因组中的岛屿。

Functional genes in the mouse and human genome are associated with CpG islands. Current estimates suggest that there may be as many as 4.5 X 10/4 islands in the human genome and slightly fewer in the mouse genome. These CpG islands are associated with both ubiquitously expressed (housekeeping) genes as well as genes with tissue-specific expression and temporal regulation. The method of restriction landmark genomic scanning (RLGS) identifies rare restriction sites in the genome as landmarks by isotopic end-labeling following enzymatic digestion. These end-labeled fragments are separated in an agarose gel following digestion with a second enzyme. The separated double-digest fragments are digested in-gel with a third restriction enzyme and the resulting products are separated in a final dimension in acrylamide. Genetic variation for these landmarks has been identified between inbred strains of laboratory mice and between common laboratory strains and different Mus species. Variation in landmarks can be mapped in the mouse genome using either standard crosses or recombinant inbred strain analyses. Genetic variation can be used to follow parent-specific alleles in other experimental circumstances. For example, methylation of CpG islands has been associated with the phenomenon of genomic imprinting that results in the epigenetic modification of parent-specific alleles for different genes that leads to a monoparental expression. Parent-specific effects are also important in the analysis of genomic instability associated with tumorigenesis. Loss- of-heterozygosity (LOH) is associated with several kinds of tumors. A simple method of scanning the genome for these kinds of events is an important experimental application of genome analysis. In a similar fashion, genetic variation for these landmarks can be used as markers for the genetic analysis of complex or multilocus traits. In this case, mice from segregating crosses showing specific phenotypes can be pooled for analysis to reduce the experimental effort for identifying genetic differences that are common to the affected progeny. Finally, it is possible to use multiple enzyme combinations of RLGS analyses for genomic regions such as congenic strains to identify candidate genes for a chromosomal region. This is a revised application of a previous proposal to genetically map RLGS loci in the mouse genome. Our primary goal in this proposal is to complete the initial stages of importing the RLGS method into our laboratory from Japan and to establish this as an experimental method for analyzing landmarks that are associated with CpG islands in the genome.