Alpha Lifetech Inc. focuses on recombinant protein expression in different Biosystems for many years, including prokaryotic and eukaryotic organisms, as well as the development of novel high-potency recombinant proteins to meet customers' requirements.
E. coli expression system, one of the prokaryotic systems of protein expression, is a famous and convenient protein expression system, known for its competitive price and high productivity for producing proteins. The basic procedures included in the process of protein expression are as follows:
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Custom Service & Procedures:
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Phase: Procedures |
Time |
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I. Gene Analysis and Synthesis |
2 weeks |
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-- Codon Optimization
-- Amplification/Isolation of target gene
-- Subcloned target gene into bacterial expression vector
-- Verification of the authenticity of the subcloned gene by restriction enzyme digest and sequencing. |
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II. Transfection & Expression |
1 week |
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-- Transfection of recombinant constructs into high-efficiency expression bacterial strain.
-- Expression optimization: induction time, temperature and IPTG concentration and so on.
-- SDS-PAGE, Coomassie blue staining and Western blot with tag-specific or custom-supplied antibody.
-- Silver staining (option) |
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III. Up-Scale Culture |
2 days |
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-- 1L of bacterial culture will be induced with inducer and harvested for protein purification. |
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IV. Purification |
1 week |
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-- Affinity purification: Ni-NTA (for 6 x HIS-tagged proteins); glutathione (for GST-tagged proteins) column
-- Non-affinity purification: IEX
-- Native or denatured condition |
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V. Endotoxin Control & Tag-free |
4 days |
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VI. QC Control |
3 days |
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-- SDS-PAGE, Western Blot or MS report |
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Total |
5 weeks |
Service Features:
-- High Success Rate: more than 95%
-- Fusion-Tag Selection: Single Tag or Combined Tags
-- Supernatant Protein Expression: Matrix Screening (over 48 conditional Permutation and combination)
-- Fermentation Advantage: Various Scales (1, 5, 10, 30, 80 L fermenter)
-- Time & Products: Short Time Cycle; High purity (no less than 95%)
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