- 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
- PURExpress Solution A
- PURExpress Solution B (Minus RF123)
- Control (DHFR) template (10 μl)
- Solution B (75 μl)
The following reagents are supplied with this product:
|Store at (°C)||Concentration|
|PURExpress Solution A|
|PURExpress Solution B|
|Control (DHFR) template|
|PURExpress Solution B|
Advantages and Features
- Quickly generate analytical amounts of protein for further characterization
- Confirmation of open reading frames
- Examination of the effects of mutations on ORFs
- Generation of truncated proteins to identify active domains and functional residues
- Introduction of modified, unnatural or labeled amino acids
- Epitope mapping
- Expression of toxic proteins
- Ribosome display
- Translation and/or protein folding studies
- In vitro compartmentalization
Properties and Usage
LicensesPURExpress® is based on the PURE System Technology originally developed by Dr. Takuya Ueda at the University of Tokyo and commercialized as the PURESYSTEM® by BioComber (Tokyo, Japan).
Licensed from BioComber (Tokyo, Japan) under Patent Nos. 7,118,883; WO2005-105994 and JP2006-340694. For research use only. Commercial use of PURExpress® In vitro Protein Synthesis Kit requires a license from New England Biolabs, Inc. This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals. Patent Nos. 7,118,883; WO2005-105994 and JP2006-340694
- The DHFR control template is now supplied at 125 ng/µl. Use 2 µl for the positive control reaction. Template DNA, particularly plasmid DNA prepared by mini-prep (e.g. Qiagen) is often the major source of RNase contamination. We strongly recommend adding 20 units Murine RNase Inhibitor (NEB #M0314) to each reaction.
- PURExpress DHFR Control Template sequence files: Fasta GenBank
- Storage: All kit components should be stored at -80°C.
- Add Solution B to Solution A, do not dilute Solution B unbuffered. We recommend a starting amount of 250 ng template DNA per 25 μl reaction. The optimal amount of input DNA can be determined by setting up multiple reactions and titrating the amount of template DNA added to the reaction. Typically, the optimal amount will fall in a range of 25–1000 ng template per 25 μl reaction.
- Asahara, H. and Chong, S. (2010). In vitro genetic reconstruction of bacterial transcription initiation by coupled synthesis and detection of RNA polymerase holoenzyme. Nuc. Acid. Res.
- Noto, T., Kurth, H., Kataoka, K., Aronica, L., DeSouza, L., Siu, K., Pearlman, R., Gorovsky, M. and Mochizuki, K. (2010). The tetrahymena argonaute-binding protein Giw1p directs a mature argonaute-siRNA complex to the nucleus. Cell. 140, 692-703.
- Tanner, D., Cariello, D., Woolstenhulme, C., Broadbent, M. and Buskirk, A. (2009). Genetic identification of nascent peptides that induce ribosome stalling. J. Biol. Chem. 284, 34809-34818.
- Talabot-Ayer, D., Lamacchia, C., Gabay, C., and Palmer, G. (2009). Interleukin-33 is biologically active independently of Caspase-1 cleavage. J. Biol. Chem. 284, 19420-19426.
- Feng, Y. and Cronan, J. E. (2009). A new member of the Eschericia coli fad regulon: transcriptional regulation of fadM (ybaW). J. Bacteriol. 191, 6320-6328.
- Solaroli, N., Panayiotou, C., Johansson, M., and Karlsson, A. (2009). Identification of two active functional domains of human adenylate kinase 5. FEBS Lett. 283, 2872-2876.
- Arenz, Stefan, Haripriya Ramu, Pulkit Gupta, Otto Berninghausen, Roland Beckmann, Nora Vázquez-Laslop, Alexander S. Mankin, and Daniel N. Wilson (2014). Molecular basis for erythromycin-dependent ribosome stalling during translation of the ErmBL leader peptide. Nature communications. 5
- Chong, Shaorong (2014). Overview of Cell‐Free Protein Synthesis: Historic Landmarks, Commercial Systems, and Expanding Applications. Current Protocols in Molecular Biology. 16-30.
- Daugherty, Ashley B., Sridhar Govindarajan, and Stefan Lutz (2013). Improved Biocatalysts from a Synthetic Circular Permutation Library of the Flavin-Dependent Oxidoreductase Old Yellow Enzyme. Journal of the American Chemical Society . 334 (38), 14425-14432.
- Desai, Bijoy J., Yuki Goto, Alessandro Cembran, Alexander A. Fedorov, Steven C. Almo, Jiali Gao, Hiroaki Suga, and John A. Gerlt (2014). Investigating the role of a backbone to substrate hydrogen bond in OMP decarboxylase using a site-specific amide to ester substitution. Proceedings of the National Academy of Sciences. 201411772.
- Gu, Liangcai, Chao Li, John Aach, David E. Hill, Marc Vidal, and George M. Church (2014). Multiplex single-molecule interaction profiling of DNA-barcoded proteins. Nature.
- Gupta, Pulkit, Shanmugapriya Sothiselvam, Nora Vázquez-Laslop, and Alexander S. Mankin (2013). Deregulation of translation due to post-transcriptional modification of rRNA explains why erm genes are inducible. Nature communications. 4
- Kaiser, Christian M., Daniel H. Goldman, John D. Chodera, Ignacio Tinoco, and Carlos Bustamante (2011). The ribosome modulates nascent protein folding. Science. 334 (6063), 1723-1727.
- Nakagawa, So, Stephen S. Gisselbrecht, Julia M. Rogers, Daniel L. Hartl, and Martha L. Bulyk (2013). DNA-binding specificity changes in the evolution of forkhead transcription factors. Proceedings of the National Academy of Sciences. 110(30), 12349-12354.
- Ramadoss, Nitya S., John N. Alumasa, Lin Cheng, Yu Wang, Sharon Li, Benjamin S. Chambers, Hoon Chang et al (2013). Small molecule inhibitors of trans-translation have broad-spectrum antibiotic activity. Proceedings of the National Academy of Sciences. 110(25), 10282-10287.
- Rosenblum, Gabriel, and Barry S. Cooperman (2014). Engine out of the chassis: Cell-free protein synthesis and its uses. FEBS letters. 588(2), 261-268.
- Stafford, Ryan L., Marissa L. Matsumoto, Gang Yin, Qi Cai, Juan Jose Fung, Heather Stephenson, Avinash Gill et al (2014). In vitro Fab display: a cell-free system for IgG discovery. Protein Engineering Design and Selection. 27(4), 97-109.
- Tuckey, Corinna, Haruichi Asahara, Ying Zhou, and Shaorong Chong (2014). Protein Synthesis Using a Reconstituted Cell‐Free System. Current Protocols in Molecular Biology. 16-31.
- Weirauch, Matthew T., Atina Cote, Raquel Norel, Matti Annala, Yue Zhao, Todd R. Riley, Julio Saez-Rodriguez et al (2013). Evaluation of methods for modeling transcription factor sequence specificity. Nature biotechnology. 31(2), 126-134.
- When using PURExpress, I was unable to synthesize the control protein?
- When using PURExpress, I was able to synthesize the control protein, but the target sample is not present or present in low yield?
- When using PURExpress, I was able to synthesize the target protein, but full-length product is not major species?
- Detailed FAQs for PURExpress?
- Are there PURExpress citations?
- Protein Synthesis Reaction using PURExpress (E6800)
- Analysis of Synthesized Protein using PURExpress (E6800)
- Determination of Protein Synthesis Yield with PURExpress (E6800)
- Purification of Synthesized Protein using Reverse His-tag Purification
- Measurement of 35S-Methionine Incorporation by TCA Precipitation and Yield Determination using PURExpress
- Use of the PURExpress In Vitro Protein Synthesis Kit Disulfide Bond Enhancer and SHuffle Competent E coli for heterologous in vitro and in vivo cellulase expression E6800
- Using the PURExpress In Vitro ProteinSynthesis Kit for Heterologous In Vitro Expression and Functional Screening of FMN dependent Oxidoreductase Variants E6800
Thoroughly mix solutions A and B before using. Do not vortex Solution B or ribosomes, mix gently.
Solution A may have a cloudy white appearance. Add to the reaction as a uniform suspension.
Assemble the reactions in the following order on ice: Solution A, Solution B, RNAse Inhibitor, Water, Template DNA or RNA
Once reaction is assembled take time to make sure everything is thoroughly mixed by gently pipetting up and down, pulse spin and place at 37C for 2 to 4 hours.