FLOW KARYOTYPING: OPTIMIZATION AND CLINICAL EVALUATION

该项目的总体目标是开发和评估改进的 产前检测疾病相关的分析细胞学技术 细胞遗传学异常 重点将放在a）提高速度和 检测染色体DNA丢失或获得的灵敏度 并在中期染色体分析期间定量，B）检测 间期细胞中常见的细胞遗传学异常和c）发育 使用分离的胎儿细胞进行细胞遗传学分析的技术 母亲的血 这些目标将通过以下方式实现： 1)应用双变量流式细胞核型分析技术定量分析 染色体缺失和重复小至1 Mb。 在此期间的努力 项目期间将致力于进一步提高 技术，并将其应用于一系列合作研究， 表征临床或分子的细微结构畸变 生物学重要性。 2)染色体特异性荧光原位杂交技术的优化 用于可靠检测和表征数值偏差的探头 涉及染色体21、18、13、x和y;节段重复 （唐氏综合征最初为21q22.3）和缺失（唐氏综合征最初为15 q11 -13）。 Prader-Willi综合征）在中期扩散和间期核。 3)开发染色体特异性探针以支持荧光 杂交研究 全染色体DNA和ssRNA探针耗尽， 重复序列将从染色体特异性粘粒产生 图书馆. 21q22.3，15 q11 -13和其他地方的复合探针将被 通过从已经映射到这些区域的“种子”探针开始生成 并在染色体特异性粘粒文库中向外“行走”， 复合探针覆盖的区域。 4)继续努力从母体血液中分离胎儿细胞， 间期细胞遗传学分析 具体项目包括： 在第一个三个月内确定胎龄， 胎儿细胞以最高频率存在于母体中， 血液，B）确定胎儿细胞（例如白细胞或 c）开发富集细胞滋养层的程序， 胎儿细胞

The overall objective of this project is to develop and evaluate improved analytical cytologic techniques for prenatal detection of disease-linked cytogenetic abnormalities. Emphasis will be on a) improving the speed and sensitivity with which the loss or gain of chromosomal DNA can be detected and quantified during analysis of metaphase chromosomes, b) detection of common cytogenetic abnormalities in interphase cells and c) developing techniques for cytogenetic analysis using fetal cells isolated from maternal blood. These objectives will be pursued by: 1) Applying bivariate flow karyotyping to quantification of the extent of chromosome deletions and duplications as small as 1 Mb. Effort during this project period will be devoted to further increasing the sensitivity of the technique and applying it in a series of collaborative studies to characterize subtle structural aberrations of clinical or molecular biological importance. 2) Optimizing fluorescence in situ hybridization with chromosome specific probes for reliable detection and characterization of numerical aberrations involving chromosomes 21, 18, 13, x and y; segmental duplications (initially 21q22.3 for Down syndrome) and deletions (initially 15q11-13 for Prader-Willi syndrome) in metaphase spreads and in interphase nuclei. 3) Developing chromosome-specific probes to support the fluorescence hybridization studies. Whole-chromosome DNA and ssRNA probes depleted in repetitive sequences will be generated from chromosome-specific cosmid libraries. Composite probes for 21q22.3, 15q11-13 and elsewhere will be generated by starting with "seed" probes already mapped to these regions and "walking" outward in chromosome-specific cosmid libraries to expand the regions covered by the composite probes. 4) Continuing efforts to isolate fetal cells from maternal blood for interphase cytogenetic analysis. Specific projects include: a) determination of the gestational age within the first trimester at which the fetal cells are present at highest frequency in maternal blood,b)determination of the origin of the fetal cells (e.g. leukocytes or cytotrophoblasts) and c) development of procedures for enrichment of the fetal cells.