Screen for molecules inhibiting histone pre-mRNA process

描述（由申请人提供）：我们将开发一种基于细胞的检测方法，使我们能够检测处理组蛋白前mRNA能力的变化。报告基因编码组蛋白mRNA的一部分，包括融合到下游GFP开放阅读框的3'端形成信号，然后是聚腺苷化信号。组蛋白3'端加工失败将导致读取到聚腺苷化位点，从而产生GFP。我们已经成功地使用这种方法进行了全基因组RNA干扰筛选。我们将调整筛选以筛选化合物。使用一个反义寡核苷酸阻断U7 snRNP，我们已经证明，我们可以很容易地检测组蛋白前mrna加工的抑制通过荧光报告。我们正在将屏幕调整为可以快速读取384孔板的自动显微镜。

DESCRIPTION (provided by applicant): We will develop a cell-based assay that allows us to detect changes in the ability to process histone pre- mRNA. The reporter encodes a portion of a histone mRNA including the 3' end formation signal fused to a downstream GFP open reading frame followed by a polyadenylation signal. Failure to process at the histone 3' end will result in read-through to the polyadenylation site resulting in production of GFP. We have successfully carried out a genome-wide RNA interference screen using this approach. We will adapt the screen to screening for chemical compounds. Using an antisense oligonucleotide which blocks U7 snRNP, we have shown that we can readily detect the inhibition of histone pre-mRNA processing by fluorescence of the reporter. We are adapting the screen to an automated microscope which can rapidly read the 384 well plates.