Step1  E. coli expression strain screening

Amplified and extracted the constructed expression plasmid.
The expression plasmids were transformed into the E. coli high expression strain.
Selection of at least five positive clones were amplified and induced to express the target protein in

LB medium. Expression after SDS-PAGE electrophoresis culture of the target protein and monoclonal

strains selected for the target protein expression appropriate. Available expression strains: DH5α,

Top10, JM115, BL21 (DE3), BL21 (DE3) pLys, XL1 Blue, XL10 Golden,Rosetta-gami.

Step 2  protein expression and purification of the target:
1L flask culture of recombinant bacteria, induce the expression of the target protein.
Through a variety of affinity chromatography, ion exchange, hydrophobic and gel filtration purification

of the recombinant protein. The removal of unwanted by protease purification tag (Tag), and then

purified to obtain the desired target protein (optional, must be added to construct an expression vector
Suitable protease cleavage sites). By SDS-PAGE electrophoresis and the results of each step to

control the final product quality. Verify the correctness of the target protein by Western-blot. Provide

customer: good target protein purification and purification 5 ~ 10mg report. Customers can request

containing purified label (Tag) or the removal of purification Label (Tag) final protein purity of up to 95%.
Time: 2 to 3 weeks

1 If you did not get the target protein , no charge. If the resulting product does not meet the requirements

of the contract and the client agrees Acceptance of the product , the costs of both parties in this step

( to give customers 20% to 50% discount ) .

2. If the customer requires detailed purification process , including detailed purification conditions and

results of each step , you need to be charged 100% of the corresponding costs .

3. Because the expression of recombinant proteins and purification of each yield is not the same,we will

ultimately provide customers with the actual situation at least 5mg,Up to 10mg of the target protein, the

charges are the same.

4.we provide a final product is generally stored in a conventional buffer . (Tris-HCl, PBS or acetic acid

- sodium acetate buffer ) protein Solution, and which does not contain urea or guanidine hydrochloride

denaturant .But because we can not build a custom activity detection system for each protein ,Therefore,

we can not guarantee that the biological activity of the final product , but we will promise to protect its

activity in accordance with the maximum possible way to prepare projects Labeled proteins. If the

customer requires a certain target protein activity , we can first provide about 50μg purified samples for

customers to detect activity. If the sample Activity substandard products , we no longer provide a target

protein to customers and received only 30% of the cost of this step , and if the sample activity pass ,

we Contract requirements will provide the same quality of the target protein and need to be charged to

the customer 100% of the cost of this step .

5 . Every Western-blot detection of up to seven samples , because you want to add a positive control,

negative control and Marker. This step contains only one Free testing , if more time is required to detect

any additional charges . We can provide customers with free anti- His-tag of an antibody, if the customer

needs To use an anti , you need to provide . In addition, as Western-blot test results affected by a

number of objective factors , we do not To ensure that the test results ( false-positive result may occur

or false negative ) . If the result is not normal, we can free redo .Steps reprocessing and detailed

detection of the target protein 3 Higher requirements on the biological activity of the purified protein

refolding of the product. Endotoxin removal, sterilization, freeze-dried for further processing.

Detailed testing the quality of the target protein by HPLC, mass spectrometry and other methods. The

target gene was subcloned into a suitable expression plasmid.

Services:
A complex study: use up to 18 different refolding buffer system, a small amount of the target protein

refolding after test samples for refolding Customer testing. According to customer requirements,

according to which the complex conditions of a small amount of sample preparation refolding proteins.

Can provide specific refolding Refolding buffer formulations and processes.

2 In addition to the toxins inside
3. Filter sterilization
4  lyophilized
5. HPLC detection
6  MS detection
Provide customers: the final product after processing and related experiments and test reports.
Time: 1 to 2 weeks

Step 4 Preparation of large scale
Provide customer: a qualified target protein and service reports.