Chromosome Analysis of Single Cells

描述（由申请人提供）：本申请旨在为候选人提供实验室研究指导计划，以促进他作为独立临床科学家的发展。在他的博士前和博士后研究期间，候选人建立了比较基因组杂交（CGH），一种基于dna的技术，用于识别染色体异常，在常规临床细胞遗传学和癌症细胞遗传学中是一种有价值的临床诊断工具。目前的研究目的是利用该工具检测单细胞的总非整倍体。由于在单个细胞上培养和进行传统的细胞遗传学分析是不可行的，因此CGH作为鉴定单个人类细胞中染色体失衡的一种有吸引力的技术。该方法的应用将为产前诊断提供来自妊娠期间母体循环的胎儿细胞的细胞遗传学分析，并为植入前诊断提供来自2至3天大胚胎的单细胞分析。因此，长期目标是(a)提供一种无创的产前诊断方法，(b)通过仅移植非整倍体胚胎来提高体外受精的着床率并降低随后的流产率。为实现这些目标，具体目标是：1。建立常规和DNA微阵列CGH方案，该方案高度敏感，特异性和可重复性，用于利用微操作装置从细胞培养中提取的单细胞进行单个人类细胞的非整倍体分析。2. 确定单细胞CGH方案的效用，以预测二倍体和非整倍体的胎儿细胞来源于母体循环在怀孕期间。3. 评估单细胞CGH方案在因荧光原位杂交（FISH）诊断为非整倍体而丢弃的胚胎中检测非整倍体的能力。4. 评估嵌合体的真实频率：(A)由于FISH诊断为非整倍体而丢弃的胚胎和(B)由于遗传疾病的分子诊断而丢弃的胚胎。候选人的发展将得到保护的研究时间、专用的实验室空间以及部门和机构的核心设施的支持。他将在负责任的研究行为的指导下，通过他的导师的参与和承诺以及西奈山医学院的研究和知识环境，他将发展成为一名独立的研究人员。

DESCRIPTION (provided by applicant): This application is designed to provide the candidate a program of mentored laboratory research to facilitate his development as an independent clinical scientist. During the course of his pre- and post-doctoral fellowships, the candidate established comparative genomic hybridization (CGH), a DNA-based technique for identifying chromosome abnormalities, to be a valuable clinical diagnostic tool in routine clinical cytogenetics as well as in cancer cytogenetics. Current research is aimed at using this tool for detection of total aneuploidy in single cells. Since it is not feasible to culture and perform conventional cytogenetic analysis on single cells, CGH presents itself as an attractive technique for identifying chromosomal imbalances in single human cells. The application of this method would provide cytogenetic analysis of fetal cells derived from maternal circulation during pregnancy for prenatal diagnosis as well as of single cells obtained from two to three day old embryos for preimplantation diagnosis. The long term goals are therefore to (a) provide a noninvasive method of prenatal diagnosis, and (b) increase the implantation rate of IVF and decrease the ensuing miscarriage rate by transferring only non-aneuploid embryos. To achieve these goals, the specific aims are: 1. Establish both a regular and DNA microarray CGH protocol which is highly sensitive, specific, and reproducible for aneuploidy analysis of single human cells using single cells extracted from cell cultures by micromanipulation apparatus. 2. Determine the utility of the single cell CGH protocols for predicting diploidy and aneuploidy in fetal cells derived from the maternal circulation during pregnancy. 3. Assess the single cell CGH protocols for their ability to detect aneuploidy in embryos discarded because of a fluorescence in situ hybridization (FISH) diagnosis of aneuploidy. 4. Assess the true frequency of mosaicism in: (A) embryos discarded because of a FISH diagnosis of aneuploidy and (B) embryos discarded because of a molecular diagnosis of an inherited disorder. The candidate's development will be supported in his endeavors with protected research time, dedicated laboratory space, and departmental and institutional core facilities. He will be guided in the responsible conduct of research, and his development into an independent researcher will be enhanced by the involvement and commitment of his mentor and by the research and intellectual environment at Mount Sinai School of Medicine.