relation: http://d-scholarship-dev.library.pitt.edu/19905/
title: Use of transcriptional synergy to augment sensitivity of a splicing reporter assay
creator: Levinson, N
creator: Hinman, R
creator: Patil, A
creator: Stephenson, CRJ
creator: Werner, S
creator: Woo, GHC
creator: Xiao, J
creator: Wipf, P
creator: Lynch, KW
description: A primary limitation in the development and use of screens to identify factors that regulate mammalian pre-mRNA splicing has been the development of sensitive reporter assays. Alternative splicing typically involves relatively small (< 10-fold) changes in isoform ratios. Therefore, reporter constructs designed to allow direct analysis of isoform expression historically have at most a 10-fold window of discrimination between a positive signal and background. Here we describe the design and application of a reporter cell line that makes use of the phenomenon of transcriptional synergy to amplify the detection of changes in splicing, such that a three- to five-fold change in splicing pattern is observed as a 30- to 50-fold change in GFP expression. Using this cell line we have identified two small molecules, from a library of approximately 300 synthetic compounds, that can induce partial repression of a variable exon from the CD45 gene. We propose that the concept of transcription-based amplification of signal will allow the development of true high-throughput screening approaches to identify effectors of mammalian alternative splicing. Copyright © 2006 RNA Society.
date: 2006-05-01
type: Article
type: PeerReviewed
format: text/plain
language: en
rights: attached
identifier: http://d-scholarship-dev.library.pitt.edu/19905/1/licence.txt
identifier:   Levinson, N and Hinman, R and Patil, A and Stephenson, CRJ and Werner, S and Woo, GHC and Xiao, J and Wipf, P and Lynch, KW  (2006) Use of transcriptional synergy to augment sensitivity of a splicing reporter assay.  RNA, 12 (5).  925 - 930.  ISSN 1355-8382     
relation: 10.1261/rna.8306