ExiProgen™ EC1 Protein Synthesis Kit

ExiProgen™ EC1 Protein Synthesis Kit is designed for synthesis and
purification of high-quality protein in under 6 hours —
simply add template DNA.

Bioneer's ExiProgen™ EC1 Protein Synthesis Kit is designed for in vitro (cell-free) expression and high-yield Histidine-Tag affinity purification of proteins by using the world's first automated protein synthesis and nucleic acid extraction system "ExiProgen™". 1 to 16 different types of highly pure proteins can be obtained from each run. Each reaction needs only 6-10 µg of DNA, which can be in the form of an expression vector such as Bioneer's pBIVT or a PCR product using the ExiProgen™ ProXpress PCR Template Kit. Because the entire process is automated, you will obtain reliable and reproducible results run after run.

Principle of protein synthesis and purification

ExiProgen™ EC1 Protein Synthesis Kit contains optimized reagents for efficient expression of various protein types, and also contains reagents and Ni-NTA magnetic beads for purification of expressed proteins. Target proteins synthesized from recombinant plasmid DNAs are bound to Ni-NTA magnetic beads and washed with washing buffer for highly pure proteins.

Features and Benefits

Fully automatic: DNA-in-Protein-out
Input 6~10ug of template DNA and load the kit into ExiProgen™ for high-purity proteins within 6 hours.

Parallel Process
Obtain from 1 to 16 different proteins in a single run.

High performance
The system allows for synthesis of proteins that are toxic to in vivo expression systems.

High purity and High yield
Each reaction well will yield up to 100ug protein that is over 90% pure.

Reproducibility
The only hands-on step is the addition of template DNA, maximizing reproducibility.

Fast process
The in vitro (cell-free) protein synthesis system does not require a time-consuming cell culture process.
 

Principle of Protein Synthesis with ExiProgen™

This product is intended to be used with Bioneer's automated system ExiProgen™ for expression and purification of proteins, and the principle of the product is illustrated in the following figure.

Left panel: Manual method of protein expression/purification – sequential and tedious
Right panel: Bioneer's automated method of protein expression/purification – simply add DNA and press start!

Template DNA

The template DNA for expression by ExiProgen™ EC1 Protein Synthesis Kit must have a "T7 promoter – ribosomal biding site (RBS) – target gene – T7 terminator" structure, and target gene must contain a histidine tag coding sequence at N-terminus or C-terminus for the protein purification.

Maximization of Protein Expression

Protein can be normally synthesized with 6-10 µg of template DNA, but the yield of protein synthesized in the reaction depends on the amount of DNA used for expression. Therefore, a preliminary experiment described as below is recommended to determine the amount of template DNA to maximize the expression of each protein using the AccuRapid™ Cell-free Protein Expression Kit. Note that more DNA is not always better!

 

Example: Synthesis of poly(A) polymerase

1. Determination of the optimal amount of template DNA for the expression of poly(A) polymerase using AccuRapid™ Cell-Free Protein Expression Kit
; Poly(A) polymerase was maximally expressed with the AccuRapid™ kit when 400 ng of pET-poly(A) polymerase was used.


Lane M: AccuLadder™ Protein Size Marker (Broad), Lane NC: Negative Control, Lane PC: Positive Control (CAT), Lane 1 - 6: 50, 100, 200, 400, 600, and 800 (ng) of pET-poly(A) polymerase

2. Protein synthesis using ExiProgen™ EC1 Proteins Synthesis Kit
6.7 ug of pET-poly(A) polymerase was used to synthesize poly(A) polymerase with the EC1 kit (400 ng x 16.7 = 6.7 ug)

Lane M: AccuLadder™ Protein Size Marker (Low)
Lane E: Expression Sample
Lane P: Purification Sample

 

Kit Contents and Storage Condition

This product contains a "Purification Kit (Kit 1)" and an "Expression Kit (Kit 2)". A Cell Extract (8-well strip tube) containing T7 RNA Polymerase and ribosomes is included separately. Cartridge ② contains a master-mix containing reagents necessary for RNA synthesis and protein expression, and Cartridge ① contains Ni-NTA magnetic beads and other buffers necessary for purification of His-tagged proteins. (The Ni-NTA magnetic beads are provided in a 20% ethanol at 10% (w/v) concentration.)

Experimental Data

1. Reproducibility: ExiProgen™ EC1 Protein Synthesis Kits provide reproducible results, the first time and every time.

Figure 1. Expression and purification of CAT. Reproducibility is seen in all 16 wells with no detectable variation between wells.
Lane 1~16: Number of the well,
M : AccuLadder™ Protein Size Marker (Low),
E : Expression sample,
P : Purification sample
 

2. Flexibility: Proteins as small as 10 kD and as large as 120 kD have been expressed/purified with ExiProgen™.


Figure 2. Expression and purification of various proteins. Up to 16 types of proteins can be expressed and purified simultaneously with an average of over 90% purity.
Top of the lane: Name of the protein,
M : AccuLadder™ Protein Size Marker (Low),
E : Expression sample,
P : Purification sample
 

3. Control: Each kit comes with a Green Fluorescent Protein positive control providing confidence in the results.

Figure 3. Detection of fluorescence emitted from fluorescent proteins synthesized with ExiProgen™.
This result indicates that these synthesized SfGFP, RFP, and AcGFP are functionally active.
Left panel: Color of each protein elution samples detected with naked eyes.
Right panel: Fluorescence from protein elution samples detected with UV illuminator.


Figure 4. Expression and purification of fluorescent proteins.
Top of the lane: Protein name,
Lane M: AccuLadder™ Protein Size Marker (Broad),
Lane E: Expression sample,
Lane P: Purification sample
 

4. Functionality: Proteins expressed/purified with ExiProgen™ can be assayed for functionality and the resulting proteins will generally yield up to 100 µg or > 90% pure protein. Many established recombinant proteins can be expressed in E. coli extracts with full functional activity.

Figure 5. Expression and purification of DUSP3 (protein tyrosine phosphatase).
Lane M: AccuLadder™ Protein Size Marker (Low),
Lane E: Expression sample,
Lane P: Purification sample


Figure 4. DUSP3 synthesized with ExiProgen™ has an enzyme activity.
TDUSP3 phosphatase activity was measured by incubating with 500 uM 3-O-methylfluorescein phosphate (OMFP).
(Reaction buffer : 100 mM Tris-HCl (pH8.2), 40 mM NaCl, 1 mM DTT, 20% Glycerol)

Application Note

• ExiProgen™_Application_Fluorescent proteins

• ExiProgen™_Application_High molecular weight protein

• ExiProgen™_Application_in vivo protein expression pET vectors

Manual

• ExiProgen™ EC1 Protein Synthesis Kit

Quality Assurance

Bioneer is the holder of Quality Management System Certificates for the following standards.

• ISO 9001:2008 - certificate

• EN ISO 13485:2003 / AC:2009 - certificate

 

FAQ
 

Q1. How long does it take to obtain purified proteins after adding template DNA to the reaction well of the kit?

Q2. What kind of template DNA is used for ExiProgen™ EC1 Protein Synthesis Kit?

Q3. How much template DNA is used for each protein synthesis reaction?

Q4. Does the purity of template DNA and codon optimization affect protein synthesis?

Q5. How much protein can be obtained using this kit?

Q6. What size of protein can be synthesized with this kit?

Q7. What is the storage temperature for this kit?

 

 

Cat.No.	Product Description	Price	Add to Cart
K-7300	ExiProgen™ EC1 Protein Synthesis Kit, 16 reactions	Inquire
K-7301	ExiProgen™ EC1 Protein Synthesis Kit, 32 reactions	Inquire
K-7302	ExiProgen™ EC1 Protein Synthesis Kit, 96 reactions	Inquire