Two Luciferase Translational Reporter System (2LUCTRS) for Assaying Translational Recoding, Reinitiation, and Internal Initiation
This invention comprises a pair of gene reporter systems that allow quantification of translational recoding, reinitiation and internal initiation of eukaryotes and prokaryotes by cloning corresponding sequences between two independent light-emitting reporters.
The invention is configured for measuring expression of two different luciferases. The system also contains a polylinker for insertion of selected genes to be tested. Recoding is determined by monitoring luminescence of one luciferase (firefly) to the luminescence of the other luciferase (renilla; i.e., sea pansy).
Some genes, including crucial genes from retroviruses such as HIV, use alteration of readout of the genetic code to set the level of their expression. This alteration is called recoding. Messenger RNA from these genes carries a special set of instructions that program ribosomes to alter the rules of readout. There is considerable interest in understanding these mechanisms, especially given that interference with them can impede viral replication. Convenient, quantitative assays are therefore helpful scientifically and for screening for molecules that inhibit or stimulate recoding.
This invention�s Dual-LuciferaseTM Reporter (DLAR) assay presents an important simplification in quantifying the expression ratios of two related translational reading frames. The invention allows automated selections for natural or synthetic substances, having enhancing or inhibiting effects on translational regulation.
Stage of Development
U.S. Patent 6,143,502 has been issued for this invention. The invention is available for further developmental research support and licensing under either exclusive or non-exclusive terms.
*Grentzmann G, Gesteland RF, Atkins JF et al. A dual-luciferase reporter system for studying recoding signals. RNA 1998 Apr;4(4):479-86.
*Wills NM, Gesteland RF, Atkins JF et al. A functional -1 ribosomal frameshift signal in the human paraneoplastic Ma3 gene. J Biol Chem 2006 Mar 17; 281(11):7082-8.
Guido Grentzmann, Raymond Gesteland, John Atkins
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