QRT-PCR TRANSCRIPT ANALYSIS

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 参与小鼠细胞糖基化和识别的各种酶和蛋白质的转录水平正在通过实时定量RT-PCR(qRT-PCR)进行定量，这种方法允许中等吞吐量(~800个基因)的转录分析。一个统一的引物设计策略正在与mRNA分离、DNA合成和qRT-PCR的优化策略相结合，这将允许在丰度为7个数量级的范围内高效地检测和测量与糖相关的基因的转录本。该方法正被用于测量来自小鼠和人类胚胎干细胞(ES)群体以及来自小鼠ES细胞的分化细胞群体的“与糖相关的”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.