A rapid method for gene expression analysis, PURExpress® is a novel cell-free transcription/translation system reconstituted from the purified components necessary for E. coli translation. With minimal nuclease and protease activity, the PURExpress system preserves the integrity of DNA and RNA templates/complexes and results in proteins that are free of modification and degradation. Transcription and translation are carried out in a one-step reaction, and require the mixing of only two tubes. With results available in a few hours, PURExpress saves valuable laboratory time and is ideal for high throughput technologies.PURExpress CitationsFigure 1: Protein expression using the PURExpress® In Vitro Protein Synthesis Kit25 μl reactions containing 250 ng template DNA and 20 units RNase Inhibitor were incubated at 37°C for 2 hours. 2.5 μl of each reaction was analyzed by SDS-PAGE using a 10–20% Tris-glycine gel. The red dot indicates the protein of interest. Marker M is the Protein Ladder (NEB #P7703, discontinued and replaced with NEB #P7717).Figure 2: Incorporation of 35S-methionine enables visualizationof protein by autoradiography25 μl reactions containing 250 ng template DNA, 20 units RNase Inhibitor and 2 μl 35S-met were incubated at 37°C for 2 hours. 2.5 μl of each reaction was analyzed by SDS-PAGE, the gel was fixed for 10 minutes, dried for 2 hours at 80°C and exposed to x-ray film for 5 hours at -80°C.Figure 3: Schematic diagram of protein synthesis and purification by PURExpressFigure 4: Expression and reverse purification of DHFR (A) and T4 DNA Ligase (B) using PURExpress125 μl reactions were carried out according to recommendations in the accompanying manual. Samples were analyzed on a 10–20% Tris-glycine gel and stained with Coomassie Blue. Note that in both cases, the desired protein can be visualized in the total protein fraction. The red dot indicates the protein of interest. Marker M is the Protein Ladder (NEB #P7703, discontinued and replaced with NEB #P7717 ).
Highlights
Cleaner System - sample degradation eliminated
Easy-to-use - protein expression complete in approximately two hours
Simple Analysis - protein can often be visualized directly on a Coomassie stained gel
This product is related to the following categories:
PURExpress,
Cell-Free Protein Expression,
Protein Expression
This product can be used in the following applications:
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