title: pDsRed-EGFPmtag-, an effective dual fluorescent reporter system for cell-based screens of premature termination codon.
abstract: A large number of inherited diseases are caused by premature termination codon (PTC) mutations that lead to the degradation of mRNA template. In this report, we developed a dual fluorescent reporter that relied the feature of fluorescent protein coding region to express a fusion protein from pDsRed-EGFPmtag-. Expression of the fusion protein from a single reporter provides a sensitive approach for high-throughput screening of cell-specific PTC events in mixed cell cultures. Results from the read-through analysis of COS7 cells carrying the nonsense mutation pDsRed-EGFPmtag-Y445X treated by PTC 124 showed EGFP transcript level was increased in the COS7 cells treated by PTC124 in a dose-dependent manner. This novel reporter system was applicable to the majority of different PTC patterns and could be used to quantify efficiency of read-through within a single cell or select cells carrying PTC.
authors.affiliation: Key Laboratory of Chemical Biology and Molecular Engineering of Ministry of Education, Institute of Biotechnology, Shanxi University, Taiyuan, 030006, People's Republic of China.
authors.forename: Quan
authors.initials: Q
authors.lastname: Shen
authors.affiliation: Key Laboratory of Chemical Biology and Molecular Engineering of Ministry of Education, Institute of Biotechnology, Shanxi University, Taiyuan, 030006, People's Republic of China.
authors.forename: Ping
authors.initials: P
authors.lastname: Guo
authors.affiliation: Key Laboratory of Chemical Biology and Molecular Engineering of Ministry of Education, Institute of Biotechnology, Shanxi University, Taiyuan, 030006, People's Republic of China. bfchai@sxu.edu.cn.
authors.forename: Baofeng
authors.initials: B
authors.lastname: Chai
keywords: Dual fluorescent reporter; Premature termination codon (PTC); Read-through efficiency