High-Throughput DNA Microarrays for Inflammation

描述(申请人提供)：一种定量高通量基因芯片 基于核酸酶保护的药物分析方法ArrayPlateTM已被开发出来 发现号。我们将利用这项技术开发两种新的分析方法，旨在 筛选影响转录因子和转录因子表达的化合物 参与炎症反应的受调控基因。对任何对象的表达式进行微小更改 其中的基因将被量化，以确定暴露于可能的候选药物的细胞。 一种检测方法将量化两种特定的转录因子，这些转录因子涉及 炎症及其调控的几个基因(共16个)。另一个将会考验 多种与免疫相关的转录因子的表达 反应以及控制其他生理途径的因素(300 总计)。将筛选2000种新型甾体化合物的模型库。 ArrayPlateTM允许在高吞吐量中使用DNA阵列技术 精确度和精密度。细胞在核酸酶保护下裂解。 寡核苷酸，它与所有靶向的mRNA杂交并保护它们。之后 核酸酶消化寡核苷酸与靶标的化学计量结合 序列在基因芯片上被量化。证实了杂交的特异性 在每个样本中。该方法足够灵敏，可以测量非常罕见的mRNA- 从在96孔板中培养的30,000个细胞降到每个细胞一个信使核糖核酸分子。

DESCRIPTION (provided by applicant): A quantitative high-throughput gene chip assay, ArrayPlateTM, based on nuclease protection has been developed for drug discovery. We will use this technology to develop two novel assays designed to screen for compounds that affect the expression of transcription factors and regulated genes involved in inflammation. Small changes in expression for any of the genes will be quantified for cells exposed to putative drug candidates. One assay will quantify two specific transcription factors involved in inflammation and several genes they regulate (16 total). Another will test expression of a wide panel of transcription factors implicated in the immune response alongside factors controlling other physiological pathways (300 total). A model library of 2000 novel steroidal compounds will be screened. ArrayPlateTM allows DNA array technology to be used in high throughput - with accuracy and precision. Cells are lysed in the presence of nuclease protection oligonucleotides, which hybridize to and protect all targeted mRNAs. After nuclease digestion oligonucleotides bound stochiometrically to targeted sequences are quantified on gene chips. Hybridization specificity is confirmed within each sample. The method is sensitive enough to measure very rare mRNAs - down to one mRNA molecule per cell, from 30,000 cells grown in a 96-well plate.