ExiProgen™ EC-Tagfree Protein Synthesis Kit

This kit is used for generating a protein without His-tag sequence with high purity on ExiProgen™. It first synthesizes a protein with a His-tag in ExiProgen™ in an automated manner. Then the His-tag is cleaved off, resulting in a pure protein without a His-tag sequence. Using protocol No. 904 the target protein can be synthesized with a high purity. All processes, including protein synthesis and dialysis, are performed in an automated manner and the final protein is harvested in any storage buffer of choice.

Features and Benefits

Protein synthesis without His-tag
A target protein is produced in a fully automated way and the His-tag is cleaved from the final protein.

Fully automated: DNA-in-Protein (in storage buffer)-out
Purified target protein will be in any storage buffer of choice.

Parallel processing and high purity
1-8 different proteins can be synthesized in a single run with purity >95%.

This kit is for an automated expression of a target protein using patented SECF (Stepwise Exchange Cell-Free) technology. The expressed protein is bound to Ni-NTA magnetic beads and then released by cleaving with TEV (Tobacco Etch Virus) Protease.

The structure of template DNA

The template DNA elements should be organized as follows.


The His-tag should be at 5' end of the target protein and TEV cleavage sequence should be placed in between the His-tag and the target protein.


The resulting purified protein will have only one extra amino acid (Glycine or Serine) at the N-terminus of the target protein. Any amino acid can be placed in the X position. Bioneer's positive control template DNA for this kit has Gln-Asn-Leu-Tyr-Phe-Gln-Gly.

Note) This kit may not be compatible with some proteins showing low solubility or low yield when it has extra amino acids at its N-terminus. Therefore, Bioneer recommends checking its expression properties using AccuRapid™ Cell-Free Protein Expression Kit (page 16) before using this kit.

Experimental results

1. Various proteins can be synthesized in a single run. The yield will be up to 200 ug per well.

M; AccuLadder™ Protein Size Marker (Low) 1; DUSP3 (22 kDa), 2; hGH (23 kDa), 3; CAT (24 kDa), 4; AcGFP (28 kDa),
* Amount of purification sample is 1/80 of total sample.
 

2. The target protein will have the His-tag removed

M; AccuLadder™ Protein Size Marker (Low)
1: Expression sample of pBIVT-TEV-AcGFP 2: His tag-TEV-AcGFP (His tagged sample), 3: AcGFP (His tag-free sample)

Application

The synthesis and purification of a protein without a tag is potentially useful for many applications including X-ray crystallography, therapeutic protein development, and antigen expression for antibody development.

Kit contents and storage conditions

This kit consists of two separate boxes, and its components are described in the following table.

Manual ExiProgen™ EC-Tagfree Protein Synthesis Kit

MSDS

MSDS ExiPrep™ EC-Tagfree Protein Synthesis Kit

Brochure

Brochure ExiProgen™ EC-Tagfree 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