MAPPING, GENE IDENTIFICATION AND DETECTION OF MUTATIONS IN CHROMOSOME 22

圆锥干先天性心脏病(CHD)患者的一组研究 已经被证明有22q11.2的微缺失。已删除的区域 很大，很可能编码几个相邻的基因。要开始 为了了解这一特定染色体片段的半合性 引起心脏缺陷的可见，此区域一定要小心 对基因进行分析和鉴定。为了实现这一点，我们建议将 该地区的详细物理地图，并使用此地图来隔离 重叠克隆的YAC和粘粒基因组DNA片段。脱氧核糖核酸 从选定的CHD患者子集中提取的样本将用于缩小 临界区。特别是，患者的断点 删除较少的代表将被研究以剖析出 基因(S)在该区域内可能负责圆锥主干 在这些患者中观察到畸形。将使用多种方法 从该区域中鉴定和分离出cDNA。这些方法将 包括新分离22号染色体特异基因的作图，直接 使用YAC和YAC选择映射到关键区域的cDNA 大白菜DNA序列的COSmids和计算机分析 关键区域的比例尺排序。在本文件中确定的成绩单 通过两种方法中的任一种杂交来确认和分离。 以战略为基础。因此，重要的发育调节基因将 被识别和刻画。它们在冠心病发病机制中的作用 将会被检查。

A subset of conotruncal congenital heart disease (CHD) patients studied have been shown to have microdeletions of 22q11.2. The region deleted is large and is likely to code for several contiguous genes. To begin to understand how hemizygosity of this particular chromosomal segment gives rise to the cardiac defects seen, this region must be carefully analyzed and the genes identified. To accomplish this we propose to make a detailed physical map of the region and use this map to isolate overlapping cloned genomic DNA fragments from YACs and cosmids. DNA samples from a selected subset of CHD patients will be used to narrow the critical region. In particular, the breakpoints of patients who represent with smaller deletions will be studied to dissect out the gene(s) within the region likely to be responsible for the conotruncal malformations observed in these patients. Multiple methods will be used to identify and isolate cDNAs from the region. These methods will include mapping of newly isolated chromosome 22-specific genes, directly selecting for cDNAs which map to the critical region using YACs and cosmids and computer analysis of the DNA sequence obtained from large scale sequencing of the critical region. Transcripts identified in this manner will be confirmed and isolated by either hybridization of PCR- based strategies. Thus, important developmentally regulated genes will be identified and characterized. Their role in the pathogenesis of CHD will be examined.