QRT-PCR TRANSCRIPT ANALYSIS

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 参与鼠细胞糖基化和识别的各种酶和蛋白质的转录水平通过实时定量RT-PCR（qRT-PCR）进行定量，该方法允许中等通量（约800个基因）转录分析。引物设计的统一策略正与mRNA分离、DNA合成和qRT-PCR的优化策略相结合，这将允许在7个数量级的丰度范围内高效检测和测量聚糖相关基因的转录本。该方法正被应用于测量小鼠和人胚胎干（ES）细胞群和小鼠ES细胞衍生的分化细胞群的“聚糖相关”转录本。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Transcript levels for various enzymes and proteins involved in murine cellular glycosylation and recognition are being quantitated by real-time quantitative RT-PCR (qRT-PCR) by a method that allows for medium-throughput (~800 genes) transcript analysis. A unified strategy for primer design is being combined with optimized strategies for mRNA isolation, DNA synthesis, and qRT-PCR that will allow for high efficiency detection and measurement of transcripts for glycan-related genes over a range of seven orders of magnitude in abundance. The approach is being applied for the measurement of "glycan-related" transcripts from mouse and human embryonic stem (ES) cell populations and differentiated cell populations derived from mouse ES cells.