Toe-printing analysis was performed using the RST2 template (49 (link)) as previously described (50 (link)). Cell free translation reactions, carried out in the PURExpress in vitro protein synthesis system (New England Biolabs, Ipswich, MA, USA) were supplemented with 50 μM thiostrepton or 100 μM of peptides which were added in water and samples (5 μl volume) were incubated for 15 min at 37°C prior to the primer extension phase of the procedure. The control reaction had no inhibitor. Primer extension was carried out for 15 min, after which the samples were processed as described (50 (link)).
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Toe-Printing Analysis for Protein Synthesis
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Corresponding Organization : Howard Hughes Medical Institute
Other organizations : University of Illinois at Chicago
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Variable analysis
independent variables
- 50 μM thiostrepton
- 100 μM of peptides
- Toe-printing analysis
- Cell free translation reactions carried out in the PURExpress in vitro protein synthesis system
- Primer extension carried out for 15 min
- Control reaction had no inhibitor
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