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DNA analysis of fetal nucleated erythrocytes (NRBCs) from maternal blood

对母血中胎儿有核红细胞 (NRBC) 进行 DNA 分析

基本信息

批准号：
10671027
负责人：
TAKABAYASHI haruo
金额：
$ 1.98万
依托单位：
Kanazawa Medical University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2000
项目状态：
已结题

项目摘要

The development of a real-time quantitative TaqMan PCR assay permits the high quality analysis of DNA or RNA. With the TaqMan PCR, Lo.et al. (1998) were successful in measuring the concentration of fetal DNA in maternal plasma and serum.Using ABI PRISM 7700 System (PE Applied Biosystems), we tried to detect the target DNA sequence of SRY and beta actin locus in the PEP (primer extension preamplification) products from a single cell. Single NRBCs (nucleated red blood cells) were retrieved from cord blood smear of male fetus using a micromanipulator.The detection rates of beta actin and SRY locus were 55% and 35%, respectively. Quantitative results of PEP products from single NRBCs were 0.5-59.5 (m=4.8) copies in beta actin and 2.4-278.4 (m=17.8) copies in SRY locus.Our quantitative data show the concentration of a single locus DNA amplified by PEP from a single NRBC. The advantages of PEP-TaqMan PCR system are high specificity and sensitivity.This system seems a powerful tool for fetal DNA diagnosis from maternal blood.

实时定量TaqMan PCR分析的发展允许DNA或RNA的高质量分析。利用TaqMan PCR（Lo.et al.（1998））成功地检测了孕妇血浆和血清中胎儿DNA的浓度，并利用ABI PRISM 7700系统（PE Applied Biosystems），从单个细胞中检测引物延伸预扩增（PEP）产物中SRY和β肌动蛋白基因座的靶DNA序列。用显微操作仪从男性胎儿脐带血涂片中回收单个NRBC，β-actin和SRY位点的检出率分别为55%和35%。PEP扩增产物的定量结果为β肌动蛋白0.5-59.5（m=4.8）拷贝数，SRY基因2.4-278.4（m=17.8）拷贝数。PEP-TaqMan PCR系统具有特异性强、灵敏度高的优点，是从母血中检测胎儿DNA的有力工具。