Molecular Dissection of the 22q11.2 Deletion Syndrome

22q11.2缺失综合征（22 q11 DS）是最常见的微缺失疾病。与其相关联 显著的表型和神经精神病理学，两者都是广泛可变的。在大多数 在受影响的个体中，由于四个大的基因介导的异常重组， 22 q11中22号染色体特异性低拷贝重复序列（LCRA、-B、-C和-D）。他们的规模和存在的许多 具有几乎相同序列的片段使染色体特异性LCR作为非等位基因的底物， 同源重组（NAHR）。NAHR的结果是许多基因组疾病，包括 22q11DS。LCR 22由于其结构上的缺陷而极难可靠地定位和测序。 特色目前，该区域的准确参考序列尚不存在。此外，LCR 22是 对短读段测序的不确定性使得它们的多态性和变异性的水平在总体上 人口不详。然而，利用BioNano Genomics的Irys技术对该地区进行光学测绘， 这个困难。事实上，我们初步的生物纳米光学映射数据表明，一个复杂的组织， LCRA和LCRD内的160 kb模块，包括拷贝数和方向差异。此外，一个共同的 在LCRC到LCRD区域的倒位多态性已经借助于Fiber-FISH鉴定。反演 会大大增加删除的风险值得注意的是，我们的初步数据表明，这种多态性 在非裔美国人（AAs）中不存在倒置，这最终解释了我们CHOP中AAs的相对不足- 基于22 q11 DS队列。在这项拟议的研究中，我们将使用创新的BioNano光学映射技术， 以确定这些22q11.2多态性在一般人群中的频率，并确定 他们在促进NAHR导致删除中发挥的作用。为了调查这些观察的基础 我们建议确定几个不同人群中LCRC-D倒位的患病率（CEU，非洲人， 和来自1000基因组计划的非裔美国人受试者;当地非裔美国人和22 q11 DS三人组）。我们 还将详细分析这些相同人群中与22 q11 DS相关的含LCR 22的区域， 决定其结构和变化。将定义反转多态的断点， 将开发PCR检测方法以筛选其作为NAHR的风险因素。最后，我们建议分析和 PCR扩增典型的3 Mb 22q11.2缺失的断点，由来自 22 q11 DS三人组中的先证者。通过利用由长单分子光学器件提供的增加的灵敏度， 在纳米通道阵列上作图，结合10 XGenomics全基因组序列和分子梳理， 这个建议将阐明以前未绘制的LCR 22及其周围环境的结构和变化 地区本文生成的数据和地图将提供对许多其他难以映射和排序的数据和地图的访问。 基因组区域。

The 22q11.2 deletion syndrome (22q11DS) is the most common microdeletion disorder. It is associated with significant phenotypic and neuropsychiatric pathology, both of which are widely variable. In the majority of affected individuals, the deletion occurs de novo as a result of aberrant recombination mediated by four large chromosome 22-specific low copy repeats (LCRA, -B, -C and -D) in 22q11. Their size and the presence of numerous segments with near-identical sequence render chromosome specific LCRs as substrates for non- allelic homologous recombination (NAHR). The results of NAHR are numerous genomic disorders, including the 22q11DS. LCR22s are extremely difficult to reliably map and sequence because of their structural characteristics. Currently, an accurate reference sequence for the region does not exist. Also, LCR22s are recalcitrant to short read sequencing such that the level of their polymorphism and variability in the general population is unknown. However, optical mapping of the region with BioNano Genomics’ Irys technology overcomes this difficulty. In fact, our preliminary BioNano optical mapping data suggests a complex organization of a duplicated 160kb module within LCRA and LCRD that includes copy number and orientation differences. Further, a common inversion polymorphism in the LCRC to LCRD region has been identified with the aid of fiber- FISH. The inversion appears to dramatically increase the risk for deletion. Remarkably, our preliminary data suggests this polymorphic inversion is absent in African Americans (AAs), which would finally explain the relative deficit of AAs in our CHOP- based 22q11DS cohort. In this proposed study, we will use the innovative BioNano optical mapping technology to determine the frequency of these 22q11.2 polymorphisms in the general population and determine the role they play in facilitating NAHR leading to the deletion. In order to investigate the basis of these observations we propose to determine the prevalence of the LCRC-D inversion in several different populations (CEU, African, and African American subjects from the 1000 Genomes Project; local African Americans and 22q11DS trios). We will also analyze in detail the LCR22-containing regions associated with 22q11DS in these same populations to determine their structure and variation. The breakpoints for the inversion polymorphism will be defined and a PCR assay to screen for it as a risk factor for NAHR will be developed. Finally, we propose to analyze and PCR amplify the breakpoint of the typical 3Mb 22q11.2 deletion guided by BioNano optical maps derived from probands in the 22q11DS trios. By leveraging the increased sensitivity afforded by long single molecule optical mapping on nanochannel arrays coupled with 10X Genomics whole- genome sequence and molecular combing, this proposal will elucidate the previously unmapped structure and variation of the LCR22s and surrounding regions. The data and maps generated herein will provide access to many other difficult to map and sequence genomic regions.