Journal: Journal of Virology
Article Title: Selective Translation of Eukaryotic mRNAs: Functional Molecular Analysis of GRSF-1, a Positive Regulator of Influenza Virus Protein Synthesis
Figure Lengend Snippet: GRSF-1 interacts with specific sequences within the influenza virus mRNA 5′ UTR as detected by gel mobility shift analysis. Recombinant GRSF-1 (100 ng) was incubated with radiolabeled NP-c, NPD3, NPD6, NPD9, NP12, NP14, M1, NA, PB1, and NP-B RNAs (10 5 dpm) in buffer that contained 5 mM HEPES (pH 7.6), 25 mM KCl, 2 mM MgCl 2 , 3.8% glycerol, 100 mM NaCl, 0.02 mM dithiothreitol, 2 mM GTP, and 1.5 mM ATP at 30°C for 20 min. The resulting RNA-protein complexes were resolved by 5% native PAGE and visualized by autoradiography. These data are representative of two independent experiments.
Article Snippet: The S10 lysate was supplemented with 0.0156 mg of tRNA per ml, 0.62 mM ATP, 0.037 mM GTP, 6.22 mM creatine phosphate, 0.0156 mg of creatine kinase per ml, 11.8 mM HEPES (pH 7.6), 1.24 mM dithiothreitol, 15.6 μM complete amino acid mixture (Promega), and 0.156 mM spermidine.
Techniques: Mobility Shift, Recombinant, Incubation, Clear Native PAGE, Autoradiography