Definition of chromosomal abnormalities by next generation sequencing

通过下一代测序定义染色体异常

• 批准号: 8063822
• 负责人: Nara Sobreira
• 金额: $ 3.84万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-04-01 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8063822
• 关键词: 10q21; 10q21.1; 17q; Base Pairing; Candidate Disease Gene; Chromosomal Rearrangement; Chromosome abnormality; Chromosomes; Cleidocranial Dysplasia; Clinic; Complex; Craniofacial Abnormalities; Cytogenetics; DNA Sequence Rearrangement; Detection; Development; Developmental Delay Disorders; Disease; Equilibrium; Evaluation; Family; Fluorescent in Situ Hybridization; Functional disorder; Generations; Genes; Genetic; Genetic Medicine; Genomics; Hereditary Disease; Heterogeneity; Human; Human Genetics; Hybridization Array; Institutes; Inverse Polymerase Chain Reaction; Karyotype; Lead; Mental disorders; Methods; Molecular; Molecular Abnormality; Molecular Cytogenetics; Mutation; NRG3 gene; Online Mendelian Inheritance In Man; Patients; Phenotype; Predisposition; Psychotic Disorders; Reading; Reporting; Resolution; Resources; Retinitis Pigmentosa; SNP genotyping; Schizophrenia; Series; Susceptibility Gene; Syndrome; Testing; Translocation Breakpoint; Usher Syndrome; base; campomelic dysplasia; clinical phenotype; comparative genomic hybridization; hearing impairment; interest; malformation; next generation; novel strategies; positional cloning; proband

DESCRIPTION (provided by applicant): Historically, chromosomal rearrangements have been investigated by conventional karyotype followed by arduous positional-cloning projects. More recently, molecular cytogenetic characterizations using fluorescence in situ hybridization, array comparative genomic hybridization, and SNP genotyping together with molecular methods such as inverse PCR and quantitative PCR have allowed an increasingly more precise evaluation of chromosome abnormalities with the result that some presumably balanced rearrangements have been found to include deletions, duplications, insertions or inversions. I will test a new approach: targeted genomic capture followed by next generation sequencing to rapidly and precisely define the molecular abnormalities in a set of clinically interesting cases followed in the Genetics clinics of the Johns Hopkins Institute of Genetic Medicine. 3 cases with complex dysmorphic syndromes already known to have translocations: the first (family L982181) is segregating an apparently balanced t(2;3)(p15;q12) translocation over 3 generations associated with craniofacial abnormalities; the second (patient JHU2010) has a t(2;6)(q22;p12.3) translocation and a breakpoint in the region of RUNX2 on chromosome 6p, associated with cleidocranial dysplasia (OMIM 119600); and the third (patient L08-2709) has a t(5;17)(q23.2;q24) translocation, with a breakpoint in the region upstream of the SOX9 on chromosome 17q, associated with the clinical phenotype of acampomelic campomelic dysplasia (OMIM 114290). Additionally, I will study three cases with deletions in the 10q21-q23 region that have been collected by the Valle lab as part of their ongoing interest in identification of schizophrenia susceptibility genes in this region. They include the fourth patient in my series (JHU2020), who has a 10q23.1 deletion associated with multiple anomalies and development delay; the fifth and sixth involve 2 families (JHU2000 and JHU88293) with overlapping deletions both of which inactivate PCDH15 on 10q21.1 and are associated with schizophrenia. The delineation of these complex rearrangements will lead to a better understanding of the molecular bases of their clinical phenotypes. In cases 5 and 6, the study of PCDH15 as a candidate gene for psychiatric disease is of interest because it is the only annotated gene that is deleted in both families (JHU2000 and JHU88293). Recessive truncating mutations in PCDH15 cause Usher type 1F (OMIM 602083) characterized by congenital hearing loss, vestibular dysfunction and pigmentary retinopathy. Interestingly, an early report, pre-appreciation of locus heterogeneity, suggested nearly 20% of patients with Usher syndrome develop psychosis [4]. PUBLIC HEALTH RELEVANCE: New methods have shown that chromosomal aberrations are more frequent than previously realized (e.g. CNV); and are often more complex than previously appreciated by examination, apparently balanced translocations by karyotype often have associated deletion or duplications at the margins. I am interested in taking advantage of chromosomal aberrations to identify genes responsible for human genetic disease and have been using methods such as FISH, SNP genotyping, qRT-PCR, inverse PCR, long range PCR, and others to define the exactly breakpoints of translocations and deletions but sometimes they do not allow the identification of breakpoints at the resolution of 1 base pair. I will test a new approach: targeted genomic capture followed by next generation sequencing (100 bp paired end reads) of the captured material to rapidly and precisely define the molecular abnormalities in a set of clinically interesting cases followed in the Genetics clinics of the Johns Hopkins McKusick-Nathans Institute of Genetic Medicine (IGM).

描述（由申请人提供）：从历史上看，常规核型随后是艰苦的位置下降项目研究了染色体重排。最近，使用荧光原位杂交，阵列比较基因组杂交以及SNP基因分型以及分子方法以及诸如逆PCR和定量PCR等分子方法的分子细胞遗传学特征，使得越来越精确的染色体异常评估与某些假设平衡的重新构造有关被发现包括删除，重复，插入或反转。 我将测试一种新方法：靶向基因组捕获，然后是下一代测序，以快速而精确地定义了在约翰·霍普金斯基因医学研究所的遗传诊所中的一系列临床有趣病例中的分子异常。 3例已知具有易位的复杂畸形综合征的病例：第一个（Family L982181）正在隔离与颅面异常相关的3代平衡T（2; 3）（P15; Q12）易位。第二个（患者JHU2010）具有t（2; 6）（q22; p12.3）的易位，在6p染色体上的Runx2区域中有一个断点，与Cleidocranial Dysplasia有关（OMIM 119600）；第三（患者L08-2709）具有T（5; 17）（Q23.2; Q24）易位，在Sox9上游的17q上的Sox9的区域中有一个断点，与Acampomelic Campomelic campomelic dospmomelic dospomim的临床表型有关114290）。此外，我将研究Valle Lab收集的10q21-Q23区域中有缺失的三个病例，这是其对鉴定该地区精神分裂症易感基因的持续兴趣的一部分。它们包括我系列（JHU2020）中的第四名患者，他的删除与多个异常和发育延迟相关。第五和第六涉及2个家庭（JHU2000和JHU88293），并具有重叠的缺失，这两者都在10q21.1上使PCDH15失活，并且与精神分裂症有关。 这些复杂的重排的描述将使人们更好地了解其临床表型的分子碱基。在病例5和6中，对PCDH15作为精神病的候选基因的研究引起了人们的关注，因为它是唯一在两个家族中删除的注释基因（JHU2000和JHU888293）。 PCDH15中隐性截断突变导致1F型usher（OMIM 602083），其特征是先天性听力损失，前庭功能障碍和色素性视网膜病变。有趣的是，早期的报告是基因座异质性的预先介绍，这表明近20％的usher综合征患者患有精神病[4]。 公共卫生相关性：新方法表明，染色体畸变比以前意识到的更频繁（例如CNV）；并且通常比检查更复杂，核型显然平衡的易位通常在边缘具有相关的缺失或重复。我有兴趣利用染色体畸变来识别负责人遗传疾病的基因，并一直使用FISH，SNP基因分型，QRT-PCR，Interve PCCR，远程PCR等方法来定义易位和易位的问题。删除，但有时它们不允许在1个基对的分辨率下识别断点。我将测试一种新方法：有针对性的基因组捕获，然后是捕获的材料的下一代测序（100 bp配对的末端读数），以迅速而精确地定义了约翰·霍普金斯（Johns Hopkins）的遗传诊所的一组临床有趣的病例中的分子异常McKusick-Nathans遗传医学研究所（IGM）。