Effective non-viral leader for cap-independent translation in a eukaryotic cell-free system

The 61 nt 5′‐untranslated region (5′‐UTR) of mRNA encoding for a light‐emitting protein of hydroid polyp Obelia longissima, obelin, is shown to provide a high level of cap‐independent translation of heterologous mRNAs in cell‐free translation systems based on wheat germ extracts. The inhibition of translation typically observed when excess mRNA is present or produced in a eukaryotic system (the so‐called self‐inhibition phenomenon) is found abated with mRNA constructs carrying the obelin mRNA leader. The role of the sequestration of a limiting initiation factor, probably eIF4F, in the self‐inhibition phenomenon and the possible independence of the obelin mRNA leader from eIF4F are discussed. We propose the obelin mRNA leader be used for effective cap‐independent translation in eukaryotic cell‐free systems, including combined transcription‐translation systems with uncontrolled phage polymerase‐catalyzed accumulation of mRNA. © 2004 Wiley Periodicals, Inc.