PUREfrex Product Pages

PUREfrex Product Manual and Package Inserts

Key Technical Notes

PUREfrex FAQ

PUREfrex in the Literature

Manufacturer's Site Links

Safety Data Sheets

Posters

  • General Protein Synthesis
    • Improvement of translational efficiency by N-terminal codon optimization in the reconstituted cell-free protein synthesis system [2016]
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    • Improvement of the translation efficiency of proline residue-containing proteins using the reconstituted cell-free protein synthesis system (PUREfrex) [2018] VIEW
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    • Investigation on how to synthesize active proteins by using a reconstituted cell-free protein synthesis system (PUREfrex) [2020]
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  • Disulfide Bonding
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    • Synthesis of proteins containing disulfide bonds using a reconstituted cell-free protein synthesis system (PUREfrex). [2019]
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    • Synthesis of proteins containing disulfide bonds using PUREfrex® with human protein disulfide isomerase [2018] VIEW
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    • Synthesis of functionally active proteins containing disulfide bonds using the new PURE system (PUREfrex) [2011]
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  • Synthesis of Immunoglobulins and Biologics
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    • Unique Cell-Free Protein Synthesis System, PUREfrex -Useful Platform for Protein Expression in the Development of Biologics- [2018] VIEW
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    • Preparation of labeled cyclic peptides with PUREfrex [2017]
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    • Development of a method for the synthesis of aglycosylated full-length IgG using PUREfrex [2017]
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    • Efficient in vitro expression of aglycosylated full-length IgG using a reconstituted cell-free protein synthesis system, PUREfrex 2.0 [2016]
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    • In vitro production of Fab, Toxin, and Immunotoxin using PUREfrex®2.0 with increased productivity [2015]
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    • Synthesis of functionally active proteins containing disulfide bonds using the new PURE system (PUREfrex) [2011]
Purefrex®Dashboard

PUREfrex® 2.0 mini [Entry model]

First choice for cell-free protein synthesis

Overview

PUREfrex® 2.0 mini is an entry model designed for those who are trying protein synthesis for the first time with a cell-free system.

Anyone can easily synthesize proteins by simply mixing template DNA and reaction solution.

It is the same product as PUREfrex® 2.0, but by making it smaller in volume, we have achieved an amazingly low price, making it easy to introduce for trial use. PUREfrex® 2.0 mini can perform reactions in as little as 10 µL, allowing you to efficiently obtain results even in small volumes. Furthermore, as a different feature from PUREfrex® 2.0, it also comes with “T7Pro-SD primer” necessary for preparing template DNA using PCR and “RNase-free water” used for preparing the reaction solution. T7Pro-SD primer is common primer to all proteins, including the essential T7 promoter and SD sequence in the 5’UTR of template DNA and is optimized for PUREfrex®.

Purefrex® 2.0 mini Ordering Information
Product NameCatalog NumberTotal Reaction Volume
PUREfrex® 2.0 mini GFK-PF201-0.1-EX 100 ul
First choice for cell-free protein synthesis

First choice for cell-free protein synthesis

PUREfrex 2.0 mini is an entry model designed for those who are trying protein synthesis for the first time with a cell-free system.

Recommended for those who have the following requests:

  • I have never used a “cell-free systems”, so I want an affordable kit for beginners.
  • I have never used cell-free systems, so I would like some support.
  • I would like to test in a small volume (10 µL～) whether the target protein can be synthesized.
  • I have never used a reconstituted system (PURE system), so I would like to try it.

About PUREfrex® 2.0 mini

GeneFrontier offers the following free support for those new to cell-free systems:

1) Detailed manual for PUREfrex 2.0 mini & email inquiry service
GeneFrontier is currently releasing a detailed manual for PUREfrex® 2.0 mini
The manual contains all the information you need to use PUREfrex® 2.0 mini, including how to design and prepare template DNA, how to prepare PUREfrex® reaction solution, and examples of protein synthesis experiments.
GeneFrontier also provides email-based support for inquiries for those who cannot understand the information from the manual alone.

How can I synthesize proteins of human origin?

  • How can I synthesize proteins of human origin?
  • What should I do if I cannot see the target protein band?
  • What should I do if the target protein has no activity? ...etc.
  • Please feel free to contact Gene Frontier researchers with your questions.
2) DNA Design for PUREfrex®
GeneFrontier offers a free service, "DNA Design for PUREfrex®", to optimize DNA template design for protein synthesis using PUREfrex®.
The design of template DNA is a factor that has an extremely large effect on the amount of target protein synthesized. As shown in the figure below, even with the same amino acid sequence, variations in DNA sequences can lead to different levels of protein production. This effect is independent of biological distinctions such as prokaryotic or eukaryotic. Certain DNA sequences tend to produce higher or lower yields of protein, and these trends have become increasingly apparent.
Figure 1. Effects of codons near N-terminal region on protein yield
A. Codons of N-terminal region of heavy chain (HC) of trastuzumab (Herceptin)
B. Comparison of yields from template DNA with different N-terminal codons

Considering these critical factors, DNA template design can often be a challenging and complex task (also see: 6 Tips about the template DNA Design).

Therefore, GeneFrontier offers a free template DNA design optimization service for those who use PUREfrex® kits. We will optimize the sequence of your target protein using template DNA sequence optimization software [PUREcodeTM].

<Consultation is free of charge!>
Send us the amino acid sequence of your target protein by the link below, and we will return a suitable DNA sequence based on our algorism considering important points for protein expression using PUREfrex®, such as codon optimization for whole ORF and N-terminus, UTRs, etc.
Request DNA design for PUREfrex®

2.0 vs 1.0 Performance

Protein yields with version 2.0 are generally significantly higher than with version 1.0 but the reaction components of v1.0 are more fully disclosed (see "components" table below). Version 1.0 is therefore more appropriate when the most precisely defined system is desirable.

PUREfrex Version Comparison
PUREfrex Version1.02.0/2.1
Specification
Yield of synthesized DHFR 150 ug/ml 600 ug/ml
Reaction volume required for 100ug DHFR 660 ul　(> 3 kits) 160 ul　(< 1 kit)
Yield of synthesized GFP 100 ug/ml 800 ug/ml
Reaction volume required for 100ug GFP 1000 ul　(> 4 kits) 125 ul　(< 1 kit)
Version Selection Guidance
Protein Preparation -- ++
Protein Science　(translation, protein folding, etc.) ++ --
Synthetic Biology ++ +

The SDS-PAGE gel below compares performance between 1.0 and 2.0 for several proteins. 1 ul of a standard 20ul reaction mixture was applied to each lane following a 4 hour reaction.

PUREfrex v2.0 has less contaminants 

Improved purification of PUREfrex® v2.0 components greatly reduced the levels of RNase and ß-galactosidase contaminants. In addition, lipopolysaccharide (LPS) levels are reduced to to approximately 0.1 EU per 1 µL of reaction mixture. All proteinous components of PUREfrex®2.0 have no tags for purification and detection. It allows to fuse your protein with any tag to purify the product.

PUREfrex 2.0 mini Kit Contents and Storage （Catalog Number: GFK-PF201-0.1-EX）
ReagentQuantityDescriptionStorage *1
Solution Ⅰ 50 µL Amino acids, NTPs, tRNAs and substrates for enzymes etc. -20 °C
Solution Ⅱ 5 µL Proteins in 30% glycerol buffer -20 °C or -80 °C *2
Solution Ⅲ 10 µL Ribosome (20 µM) -80 °C *2
DHFR DNA *3 10 µL PCR product containing a gene encoding dihydrofolate reductase (DHFR) from E.coli as a positive control. (20 ng/µL) -20 °C
T7PRO-SD primer 5 µL Primer common to all proteins, including the essential T7 promoter and SD sequence in the 5’UTR of template DNA (10 µM) -20 °C
RNase-Free Water 500 µL RNase-Free Water -20 °C
  • ＊1) Indicated temperature shows storage temperature after opening the kit. Please store the kit at -80 °C before opening.
  • ＊2) The rest of solution should be frozen rapidly in liquid nitrogen or dry ice/ethanol, and be stored at -80 °C. Divide into aliquots, if necessary, and avoid refreeze and thaw as much as possible.
  • ＊3) For synthesizing DHFR, add 1 µL of DHFR DNA to 20 µL of reaction. Sequence of DHFR DNA is here.

PUREfrex 2.0 mini General Reaction Protocol

  • 1)Indicated temperature shows storage temperature after opening the kit. Please store the kit at -80 °C before opening.
  • 2)Thaw Solution I by incubation at room temperature or 37 °C for 1 minute for completely dissolving, and then cool on ice.
  • 3)Thaw Solution II and III on ice.
  • 4Mix Solution I, II and III respectively by vortex and centrifuge briefly to collect each solution at the bottom.
  • 5)Assemble the reaction mixture in a tube as follows. Add the template DNA to 1 - 3 ng/µL per 1 kb.
  • 6)Incubate the tube at 37 °C for 4 - 6 hours on a heat block or a water bath.
  • 7)Analyze the synthesized products. Please add the same amount of H2O to the reaction for the sample of SDS-PAGE.
Contents of a standard PUREfrex reaction
Contents of a standard PUREfrex reactionvolume
Water 3.5-X μL
Solution I 5 μL
Solution II 0.5 μL
Solution III 1 μL
Template DNA * X μL
Total 10 μL
* Please visit the template DNA preparation site.

PUREfrex Ordering Information

PUREfrex Ordering Information
Product Name
(click for price and order info)		Catalog NumberTotal Reaction VolumeNotes
Entry Model Kits
PUREfrex® 2.0 mini GFK-PF201-0.1-EX 100 μL New item !
PUREfrex® 2.0 mini is an entry model designed for those who are trying protein synthesis for the first time with a cell-free system.Product Information Page：PUREfrex® 2.0 mini
Kits
PUREfrex® 2.0 GFK-PF201-0.25-EX 250 ul  
GFK-PF201-0.25-5-EX 5 x 250 ul (packaged as 5 x 250 ul) Sales to be discontinued
GFK-PF201-2ML-EX 1 x 2 ml  
GFK-PF201-10ML-EX 5 x 2 ml Note: cat. no. change from GFK-PF201-10-EX
GFK-PF201-50ML-EX 25 x 2 ml New size !
Current generation kit for protein synthesis. Contains DTT as a reducing agent. Product Information Page：PUREfrex® 2.0
PUREfrex® 2.1 GFK-PF213-0.25-EX 250 ul  
GFK-PF213-0.25-5-EX 5 x 250 ul (packaged as 5 x 250 ul) Sales to be discontinued
GFK-PF213-2ML-EX 1 x 2 ml New size!
GFK-PF213-10ML-EX 5 x 2 ml Note: cat. no. change from GFK-PF213-10-EX
GFK-PF213-50ML-EX 25 x 2 ml New size !
Current generation kit for proteins with disulfide bonds. Formulated for user control of redox environment. Product Information Page：PUREfrex® 2.1
PUREfrex® 1.0 GFK-PF001-0.25-EX 250 ul  
GFK-PF001-0.25-5-EX 1250 ul (packaged as 5 x 250 ul)  Sales to be discontinued
GFK-PF001-2ML-EX 1 x 2 ml  New size!
 GFK-PF001-10ML-EX 5 x 2 ml Note: cat. no. change from GFK-PF001-10-EX
 GFK-PF001-50ML-EX 25 x 2 ml New size !
Prior generation kit for protein science research and synthetic biology. Components are fully disclosed. Product Information Page：PUREfrex® 1.0
Custom formulated kits are based on 1.0 components. Please inquire about PUREfrex® custom.
PUREfrex Reaction Supplements
DnaK Mix GFK-PF003-0.5-EX for 500 ul of reaction mix  
GFK-PF003-10ML-EX 5 x 2 ml  Note: cat. no. change from GFK-PF003-10-EX
GFK-PF003-50ML-EX 25 x 2 ml New size !
GroE Mix GFK-PF004-0.5-EX for 500 ul of reaction mix  
PF004-10ML-EX 5 x 2 ml  Note: cat. no. change from GFK-PF004-10-EX
PF004-50ML-EX 25 x 2 ml New size !
Supplements for synthesis of aggregate-prone proteins. Product Information Page：PUREFrex Reaction Supplements
DsbC Set GFK-PF005-0.5-EX for 500 ul of reaction mix  
PF005-10ML-EX 5 x 2 ml Note: cat. no. change from GFK-PF005-10-EX
PF005-50ML-EX 25 x 2 ml New size !
PDI Set GFK-PF006-0.5-EX for 500 ul of reaction mix   
PF006-10ML-EX 5 x 2 ml Note: cat. no. change from GFK-PF006-10-EX
PF006-50ML-EX 25 x 2 ml New size !
Supplements for synthesis of proteins containing disufide bonds.. Product Information Page：PUREFrex Reaction Supplements
EF-P GFK-PFS052-0.5-EX for 500 ul of reaction mix   
PFS052-10ML-EX 5 x 2 ml Note: cat. no. change from GFK-PFS052-10-EX
PFS052-50ML-EX 25 x 2 ml New size !
Supplement for use when consecutive prolines are in target protein.  Product Information Page：PUREFrex Reaction Supplements

Notes

PUREfrex® is developed for in vitro research only. PUREfrex® should not be used for the therapy, diagnostic or administration to animals including human and should not be used as food or cosmetics etc. To avoid the contamination of nuclease, nuclease-free-treated water, reagents and materials should be used. We also recommend wearing gloves and mask.
“PUREfrex® is Registered in U.S. Patent and Trademark Office”