1. About PUREfrex®
PUREfrex® is a reconstituted cell-free protein synthesis#PFS052-12ML-EX
For 2 mL x 6 Reaction
PUREfrex® is NOT included.

in vitro research use only
Store at -80ºC before opening
GeneFrontier Corporation
www.genefrontier.com

2024 Summer
SHARP Kashiwa Building, 4F
273-1 Kashiwa, Kashiwa-shi,
Chiba 277-0005 Japan

EF-P *1

50 µL x 640 µM EF-P (in 30 % glycerol buffer)

Dilution BufferStore at -80ºC*2

500 µL x 1kit which GeneFrontier has developed based on the PURE
system technology. The target protein can be synthesized
by adding the template DNA (or mRNA) to the reaction
mixture. The PURE system is a unique cell-free protein
synthesis system, which has originally developed byProfessor Takuya Ueda at the University of Tokyo, andc o n s i s t s o f o n l y p u r i fi e d f a c t o r s n e c e s s a r y f o rtranscription, translation and energy regeneration (Ref. 1).
Therefore it enables to adjust the composition of thereaction mixture.
PUREfrex® has been raised in purity by improving themethods for preparing ribosomes, tRNAs and all proteinsin the reaction mixture compared with the original PURE
system (Ref. 2). As the result, the contaminating
lipopolysaccharide from E. coli is reduced to around 0.1 EU
per 1 µL of reaction and other contaminants, such as
RNase and β-galactosidase, are also reduced.
All of proteins in PUREfrex® have no tags, thesynthesized protein can be purified and detected by anytags.

2. About EF-P
EF-P (elongation factor P) is one of the translation
factors in Escherichia coli (E. coli) and a homolog ofeukaryotic initiation factor 5A (eIF5A). EF-P improves thesynthesis of protein containing consecutive proline
residues such as Pro-Pro-Pro and Pro-Pro-Gly bypromoting the formation of peptide bonds. Lysine at 34thof EF-P is post-translationally modified to β-lysillysine andthe modification is important for the activity.
Because PUREfrex® 1.0, PUREfrex® 2.0 andPUREfrex® 2.1 don't contain EF-P, the synthesis efficiency
of some proteins containing consecutive proline residues
are low. Addition of EF-P to the PUREfrex® reaction
mixture increase the productivity of such proteins.
EF-P (#PFS052-0.5) is a supplement for PUREfrex® thatcontains recombinant post-translationally modified EF-P
isolated from E. coli.

Store at lower than -20ºC*2)For storage at -80ºC, the rest of solution should be frozen rapidly
in liquid nitrogen or dry ice/ethanol. Please divide into aliquots, if
necessary, and avoid refreeze and thaw as much as possible.

【JAPAN】

Carlsbad, CA 92010, USA

KOTO-KU. TOKYO 135-0016, JAPANemail: info@cosmobiousa.com

Phone/FAX: (+1) 760-431-4600

2. Shimizu et al. (2005) Methods, vol. 36, p. 299*1)Standard final concentration of EF-P is 0.1 – 2 µM.
We recommend to check the optimal concentration of EF-P
because it depends on a protein of interest.

【Outside Japan】

2792 Loker Ave West, Suite 101References) 1. Shimizu et al. (2001) Nat. Biotecnol., vol. 19, p. 751

Store at -80 ºC before opening

EF-P and PUREfrex® is developed for in vitro researchuse only. EF-P and PUREfrex® should not be used for thetherapy, 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 andmask.
For information concerning commercial use of EF-P,
please contact GeneFrontier.
e-mail : purefrex@genefrontier.com

URL: www.cosmobiousa.com

Here is a standard protocol for synthesizing proteinsusing EF-P and PUREfrex® 2.1 (#PF213). For example,
please assemble 20 µL of reaction mixture as below, inwhich the final concentration of each reagent is 0.5 mM
Cysteine, 4 mM GSH and 1 µM EF-P.

TOYO EKIMAE BLDG. 2-20, TOYO 2-CHOME,
Phone: +81-3-5632-9610
FAX: +81-3-5632-9619

URL: https://www.cosmobio.co.jp/

5. Incubate the tube at 37ºC for 2-6 hours with heat blockor water bath.
6. Analyze the synthesized product.

1. Thaw Solution I, Cysteine and GSH by incubation atroom temperature or 37 ºC for 1 minute completely, andthen cool on ice.
2. Thaw Solution II, III and EF-P on ice.
3. Mix each solution by vortex and centrifuge briefly tocollect each solution at the bottom.
4. Assemble the reaction mixture in a tube as follows.
(Add the template DNA to 0.5-3 ng/µL per 1 kbp)

Water
Solution I *3
10 mM Cysteine
80 mM GSH
Solution II
Solution III
40 µM EF-P
Template DNATotal

6.5-X µL
8 µL
1 µL
1 µL
1 µL
2 µL
0.5 µL
X µL
20 µL*3)
Please note that the volume of Solution I of PUREfrex® 2.1 (#PF213)
is different from PUREfrex® 2.0 (#PF201).