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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]

Which PUREfrex Kit Is Right For Me?

PUREfrex® coupled transcription/translation system kits are available in four versions. 

  • PUREfrex® 2.0 - Choose 2.0 when protein yield is you primary goal and disulfide bonding is not relevant.
  • PUREfrex® 2.1 -  2.1 kits are based on 2.0 reaction components but are not premixed with reducing agent. 2.1 kits include separate vials of cysteine, DTT, and reduced glutathione allowing control of the redox environment. Choose 2.1 kits for primarily for target proteins with disulfide bonds.   
  • PUREfrex® 1.0 - 1.0 kits produce less protein than 2.X kits, but the complete contents of 1.0 kits is disclosed. Chose 1.0 when complete information on PUREfrex reaction components takes precedence over protein yield. 
  • PUREfrex® 1.0 Custom - Gene Fronteir is happy to provide custom reaction mixtures whenever feasible. Custom PUREfrex kits are based on 1.0 reaction components. Please contact us with your request.  

For detailed information about each PUREfrex® version, please use the navigation links in the sidebar on this page.

About Purefrex 1.0

The protein yield of Purefrex 1.0 is less than 2.0 and 2.1 but the the contents of 1.0 are fully disclosed (see "components" table below) making PUREfrex 1.0 the preferred version for basic research on translation, protein folding, synthetic biology, and other topics where absolute protein yield is less critical. Researchers interested in custom formulated kits should know that PUREfrex custom kits are based on version 1.0. Initial testing of your intended experimental templates with a PUREfrex 1.0 kit is therefore advisable prior to requesting custom formulations.  

 

Protein Yield of PUREfrex Kit Versions. 
PUREfrex Version 1.0 2.0/2.1
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)

 

 Otherwise, operation of PUREfrex version 1.0 kits are essentially identical to version 2.0 and are compatible with all PUREfrex supplements. For more information on PUREfrex supplements, please review the navigation sidebar on this page.

PUREfrex 1.0 Kit Contents
Reagent Quantity Description Storage
Storage Solution I 125 µL Amino acids, NTPs, tRNAs and substrates for enzymes etc.

-20 °C
Solution II 12.5 µL Proteins in 30 % glycerol buffer

-20 °C or -80 °C*2
Solution III 12.5 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

 PUREfrex version 1.0 kits contain the following components. 

PUREfrex 1.0 Reaction Components
Solution 1 (Buffers, Salts, Substrates Solution II (proteins)  Solution III
Hepes-KOH pH 7.6

20 Aminoacyl-tRNA synthetases (ARS)

 Ribosomes 
Potassium glutamate

Methionyl-tRNA transformylase (MTF)

  
Magnesium acetate

3 Initiation factors (IF1, IF2, IF3)

  
Spermidine

3 Elongation factors (EF-Tu, EF-Ts, EF-G)

  
Dithiothreitol

3 Release factors (RF1, RF2, RF3)

  
10-Formyl-tetrahydrofolate

Ribosome recycling factor (RRF)

  
4 NTPs

T7 RNA polymerase

  
20 Amino acids

Nucleoside diphosphate kinase (NDK)

  
Creatine phoshate

Adenylate kinase (ADK, myokinase)

  
tRNA mixture

Ceatine kinase (CK)

  
 

Pyrophosphatase (PPiase)

  

 

PUREfrex 1.0 General Reaction Protocol

1) The amounts given are for 20 µL reaction. For scaling up the reaction, adjust the volume of reagents accordingly.
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.
4) Mix 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 2 – 4 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.qqqqqqq
Contents of a standard PUREfrex reaction

volume
Water

8-X µL
Solution I

10 µL
Solution II

1 µL
Solution III

1 µL
Template DNA *

X µL
Total

20 µL
* Please visit the template DNA preparation site
PUREfrex Ordering Information
Product Name
(click for price and order info)		 Catalog Number Total Reaction Volume Notes Product Information Page
Kits
PUREfrex 2.0
 GFK-PF201-0.1-EX 100 ul New Size! 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   New size!
Current generation kit for protein synthesis. Contains DTT as a reducing agent.
PUREfrex 2.1
 GFK-PF213-0.25-EX 250 ul   2.1
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  
 GFK-PF213-10ML-EX  5 x 2 ml Note: cat. no. change from GFK-PF213-10-EX
GFK-PF213-50ML-EX   New size!
Current generation kit for proteins with disulfide bonds. Formulated for user control of redox environment. 
PUREfrex 1.0
 GFK-PF001-0.25-EX 250 ul   1.0
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  
 GFK-PF001-10ML-EX 5 x 2 ml Note: cat. no. change from GFK-PF001-10-EX
GFK-PF001-50ML-EX   New size!
Prior generation kit for protein science research and synthetic biology. Components are fully disclosed.
Custom formulated kits are based on 1.0 components. Please inquire about custom kits.   
PUREfrex Reaction Supplements
DnaK Mix GFK-PF003-0.5-EX for 500 ul of reaction mix   Supplements 
GFK-PF003-10ML-EX 5 x 2 ml  Note: cat. no. change from GFK-PF003-10-EX
GFK-PF003-50ML-EX   New size!
GroE Mix GFK-PF004-0.5-EX for 500 ul of reaction mix   Supplements 
PF004-10ML-EX 5 x 2 ml  Note: cat. no. change from GFK-PF004-10-EX
GFK-PF004-50ML-EX   New size!
Supplements for synthesis of aggregate-prone proteins.
DsbC Set GFK-PF005-0.5-EX for 500 ul of reaction mix   Supplements 
PF005-10ML-EX 5 x 2 ml Note: cat. no. change from GFK-PF005-10-EX
GFK-PF005-50ML-EX   New size!
PDI Set GFK-PF006-0.5-EX for 500 ul of reaction mix    Supplements 
PF006-10ML-EX 5 x 2 ml Note: cat. no. change from GFK-PF006-10-EX
GFK-PF006-50ML-EX   New size!
Supplements for synthesis of proteins containing disufide bonds.
EF-P GFK-PFS052-0.5-EX for 500 ul of reaction mix    Supplements 
PFS052-10ML-EX 5 x 2 ml Note: cat. no. change from GFK-PFS052-10-EX
GFK-PFS052-50ML-EX   New size!
Supplement for use when consecutive prolines are in target protein.

 

Custom Kit Examples

  1. (-) RF1: For synthesis of a protein containing a noncanonical amino acid by amber suppression as below.
  2. (-) T7 RNA polymerase: For use of other RNA polymerase such as E.coli RNA polymerase.
  3. (-) Specific amino acid(s) and/oraminoacyl-tRNA synthetase:  For synthesis of a protein containing noncanonical amino acid or less than 20 amino acids.

 

 

Agreement

Custom kits are sold under the agreement of GeneFrontier (“GFC”). Rights to use this product are limited.

Condition of Use

  1. Material is for internal research use only, and for non-clinical use
  2. Material is for non-commercial use, and not allowed to distribute to any third parties
  3. Material is not to be modified beyond the Purpose of Use
  4. Customer shall promptly notify GFC if any new invention is made through the use of Material or its portion
  5. Any new invention relating to Material and the use belongs to GFC
  6. Any products itself synthesized through the use of Material, and the ownership of the synthesized products belong to Customer
  7. Industrial customers need additional “License Agreement” for the use of in vitro display

 

How to order 

  1. Request an agreement and check the terms and conditions of use.
  2. Ask possibility if your designed kit could be prepared.
  3. Sign the agreement.
  4. Receive product number for your designed kit.
  5. Order with the received product number.

Request for Quotation

  • Price is set for each reaction volume.
  • The minimum reaction volume that can be ordered is 1 mL, it is sold in mL units. This is different from the unit of the kit.
  • Please request a quote to Cosmo Bio USA.

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Purefrext Supplements Overview

PUREfrex supplements add functionality to PUREfrex reaction mixtures. Adding the right supplement could mean the difference between success and failure to obtain the outcome you hope for. All supplements are prepared in buffers and at concentrations that allow their use directly in standard PUREfrex reaction protocols. Like all PUREfrex protein components, those supplements that are proteins are also untagged.  Chaperones DnaK

help 

Redox-state ModifiersDsbCPDI SetTranslation FactorsEF-P

 

 

 

 

Overview

PUREfrex® coupled transcription/translation system kits are available in four types. 

  • PUREfrex® 2.0 - use 2.0 when protein yield is you primary goal and disulfide bonding is not relevant.
  • PUREfrex® 2.1 - use 2.1 when protein disulfide bonding is needed.  
  • PUREfrex® 1.0 - use 1.0 when complete information on PUREfrex reaction components is more important than protein yield. 
  • PUREfrex® 1.0 Custom - inquire about custom kits to alter the components included in PUREfrex 1.0 reaction solutions.   

For more detailed information about PUREfrex®, click here

  

PUREfrex® 1.0 and PUREfrex® Custom Kits

For translation research, synthetic biology, mechanistic studies, etc.

PUREfrex® 1.0 - 1.0 kits are for use in basic research such as in studies on the mechanisms and regulation of of protein synthesis. Component information for PUREfrex® 1.0 is publicly disclosed- but the protein yield is lower than with PUREfrex® 2.0 and 2.1. It is important to recognize that all PUREfrex® 1.0 custom kits (e.g., kits without one or more transcription factors) are based on PUREfrex® 1.0. Before ordering a custom kit, it is recommended to verify the protein of interest can be synthesized with a PUREfrex® 1.0 kit. For more information on custom kits, please see the Custom Kits webpage.  

PUREfrex® 2.0 and PUREfrex® 2.1

To disulfide bond or not to disulfide bond? PUREfrex has an answer.  

PUREfrex® 2.0 and PUREfrex® 2.1 - If protein yield is your primary goal, chose a 2.X kit.

PUREfrex® 2.0 contains the reducing agent DTT to inhibit disulfide bonding. If you do not intend to produce protein with disulfide bond(s), chose 2.0.   

PUREfrex® 2.1 is designed specifically for synthesizing proteins with disulfide bonds. Because no one set of redox environment conditions will support support disulfide bonding for all proteins, PUREfrex® 2.1 is thoughtfully formatted for user control of the redox environment, enabling a variety of conditions to be easily evaluated.