Journal: BMC Biotechnology

Article Title: A comparative study of protein synthesis in in vitro systems: from the prokaryotic reconstituted to the eukaryotic extract-based

doi: 10.1186/1472-6750-8-58

Figure Lengend Snippet: The effect of the size-separated fractions on in vitro protein synthesis . A. Screening the fractions (10 to 21) from a size-exclusion separation of an S30 extract for enhancement in in vitro protein synthesis in the PURE system. The activities were assayed at 60 min. for Fluc (grey bars) and β-galactosidase (black bars), and by comparing to the activities of the controls (supplemented with only the column buffer), the data are presented as "fold increase". B. Comparison of the protein synthesis yields of Fluc between the PURE system (PURE) without any supplement (Buffer) or supplemented with the fraction 17 (F17) or the S30 extract (Sup), with the S30 systems from Invitrogen (Expressway) and Roche (RTS). The yields were calculated from the activities of the in vitro synthesis reactions compared to that of the known amount of pure Fluc protein assayed at the same time.

Article Snippet: This result remained the same whether the crude RRL (Pel-Freez) or the RRL from the in vitro protein synthesis kit (Promega), which presumably contained functional eukaryotic chaperones, was used as the supplement (data not shown).

Techniques: In Vitro

Journal: BMC Biotechnology

Article Title: A comparative study of protein synthesis in in vitro systems: from the prokaryotic reconstituted to the eukaryotic extract-based

doi: 10.1186/1472-6750-8-58

Figure Lengend Snippet: In vitro synthesis of the fusion protein FG . A. Western blot analyses (anti-Fluc) of the in vitro synthesis reactions (at 60 min.) of Fluc, FG and F-SH in different systems. The full-length proteins, indicated by arrows, were confirmed by using the purified proteins of Fluc (purified Fluc) and FG (purified FG). B. The Fluc activities were determined for the Fluc and FG templates after 60 min. of the in vitro synthesis reactions in different systems. The data are presented as the relative activities with the activity of Fluc in S30 at 60 min being 100. C. Western blot analyses of the time courses of the in vitro synthesis reactions of FG in S30+RRL and RRL using anti-Fluc and anti-GFP to probe the translation products. The positions of the full-length FG and Fluc, indicated by arrows, were confirmed by the purified FG (purified FG) and the in vitro synthesis reaction of Fluc (FLuc control). The molecular weights (kDa) are indicated at the left.

Article Snippet: This result remained the same whether the crude RRL (Pel-Freez) or the RRL from the in vitro protein synthesis kit (Promega), which presumably contained functional eukaryotic chaperones, was used as the supplement (data not shown).

Techniques: In Vitro, Western Blot, Purification, Activity Assay