Bacillus Megaterium High-Yield Expression Service

Creative BioMart has developed a high-yield Bacillus megaterium expression system that exhibits 10 times more productivity than the classic ones. For customers who need a large quantity of protein products, please consider this service.

Technology Description

The main difference between the high yield and the classical Bacillus megaterium expression system is the structure. We modified the basic plasmid carrying natural promoter.

The enhanced vectors include vectors carrying two different signal peptide sequences, a 6xHis-tag, a Strep-tag, or two ribosome binding sites (RBS) for simultaneous dual expression. Vectors encoding C- or N-terminal His-tags for easy purification are also offered. The protein secretion with LipA or YocH signal peptides is up to 9-fold increased. All enhanced vectors have established multiple cloning sites for versatile cloning. Induction of protein expression is achieved by the tightly regulated and efficiently inducible xylose operon.

Technology Features

High-performance vector with an optimized sequence
Ideal for small and large scale protein production
Strictly regulated and effectively induced xylose operon (up to 350 fold)
No alkaline protease activity even up to 5 hours after induction
No observed endotoxin
Multiple cloning sites can be used for multifunctional cloning
Coding C-terminal or N-terminal his tag for universal purification (NATURAL 6xHis tag)

Service Content

Below is Creative BioMart's high-yield Bacillus megaterium protein expression procedure and deliverable materials.

Bacillus Megaterium High-Yield Expression Service

1. Selection of expression system (if the customer provides the expression strain, this step will be omitted. Only the optional host strain of expression plasmid will be provided.)
Host strains and shuttle vectors selection	Contact us for the available vectors and strains
2. Shuttle vector construction	Deliverable
Gene optimization Subcloning design Synthetic gene Cloning strategies selection Subclone the GOI to shuttle vector	1. Gene synthesis report 2. Sequencing validation report
3. Obtain the recombinant shuttle vector
Recombinant shuttle vector cultured in E.coli strain Shuttle vector extraction	1. Cloning strain containing expression plasmid (optional) 2. shuttle plasmid 3. Plasmid instructions
4. Analysis and identification of protein expression test
Transform the shuttle vector into Bacillus megaterium protoplasts Expression test Expression analysis and identification Expand scale	1. Positive expression strains 2. Small test expression process report 3. Images and spectrums obtained from the analysis 4. Small test expression summary
5. Protein expression and purification
Protein affinity purification (extracellular or intracellular) Identify purified proteins using WB Ammonium sulfate precipitation of proteins in the cell-free supernatant	1. Protein purification process report 2. Original picture of protein purification 3. Delivery of protein 4. Western blot (labeled-antibody) 5. Protein fingerprint (optional) 6. Molecular weight of protein mass spectrometry (optional)
6. Scale-up culture in fermentor
Amplification Culture (10-20l) Amplification Culture (above 50L)	1. Protein purity identification report 2. Finished protein (agreed price according to the results of expression test)

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