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Exogenous Expression

Product Introduction


Using molecular biological techniques, we construct the exogenous expression vector by ligating the targeted gene to protein expression vector. Express the target proteins with E.coli expression system, yeast expression system, insect expression system and mammalian expression system. By purifying the target protein through affinity chromatography, ion exchange chromatography, gel filtration chromatography and so on, we obtained the aimed protein sample met requirements for studying the D-structure and function.

Advantages

  • Quality: We provide a “one-stop service” from gene synthesis to protein expression and purification. We can provide codon optimization analysis, using a variety of fusion label vectors and hosts to ensure high soluble protein expression and high protein activity. With almost 100 fermenters of different sizes from 1L to 500L, it can meet the requirements of high throughout and large-scale recombinant protein expression.
  • Specialty: Four systems of protein expression;Various purification systems could both purify natural protein and recombinant protein.

Project Workflow

Turnaround Time

E.coli expression system Yeast expression system Insect expression system Mammalian expression system
7-8 weeks 8-9 weeks 8-9 weeks 7-8 weeks

 

Sample Requirements

 

The protein sequence can be directly provided, or the recombinant vector.

Frequently asked questions

 

What is the purpose of codon optimization?

(1) To remove the rare codon;
(2) To optimize the secondary structure of mRNA;
(3) To optimize the ratio of GC;
(4) To change the restriction enzyme site;
(5) To improve the efficiency of translation termination.

What should be done when the protein is not expressed, the band is too weak and the target protein is insoluble or expressed as inclusion body?

Please try to change the expression system, change the expression vector, change the host cell, explore the appropriate expression conditions or even redesign the gene.

How to improve the purity of target protein?

If the purity of protein does not meet the requirements, you can try using ion exchange chromatography and gel filtration chromatography to separate and purify the protein for further improving the purity of protein.


Amplification of target gene by PCR

After gene synthesis, using high – fidelity enzyme to amplify target gene, gene is 1,065 bp in size, as the bright stripes shown in the figure. The successful amplified genes are prepared for the construction of recombinant plasmid.


Identification of Recombinant Plasmid by Double Digestion

Whether target gene is successfully ligated to the vector could be identified by the double digestion. As shown in figure 2, the specific cutting results by using restriction enzyme BamHI and XhoI to treat recombinant plasmid indicated that the gene was successfully ligated to the vector.


Protein Expression Test

Test the expressed product of target gene by using SDS-PAGE and WB and the results shown in figure 3 and 4 confirmed the target pro- tein was expressed after induction and the molecular weight is around 60 kDa.


Protein Expression Test

Purify the target protein by using Ni-NTA affinity chromatography: with the imidazole concentration increasing, the purity of target protein increased, such as the strips shown in lane 6, figure 5, the purify is more than 90%.


Affinity Chromatography Purification

Through gel filtration chromatography, the elution curve and volume of target protein could identify the aggregation form of target protein, also the elution product is high quality purified. As shown in figure 6, after purified by the gel filtration, the target protein is homogeneous monomer form in solution and the purity is over 95%.