Custom Protein Expression – Get It Your Way

Abgent's selection of bacterial and baculovirus protein expression systems provides a choice for your application. We will work with your specifications for quantity, bioactivity, structure, post-translational modifications, and purity to express proteins that meet your specialized requirements. Project status and data reports are monitored by our exclusive in-house custom service database tracking system.

        Description of Protein Expression in Bacterial System
        Description of Protein Expression in Baculoviral System
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Protein Expression in Bacterial System	Download PDF	Order Form
Phase Time I. Cloning of a gene of interest into a bacterial expression vector1 Amplification/isolation of the gene of interest out of a customer-supplied vector and subcloning it into a bacterial expression vector. Verification of the authenticity of the subcloned gene by restriction enzyme digest and sequencing. Maxi-prep of the recombinant vector DNA. 2 weeks II. Generation and identification of bacterial colonies expressing target proteins Transformation of recombinant constructs into high-efficiency expression bacterial strain. Mini-induction to over-express the target protein. Test for the expression of the recombinant protein by western blot (customer must provide appropriate antibodies) if desired. 1 week III. Large-Scale Culture One liter of bacterial culture will be induced with IPTG and harvested for protein purification. 2 days IV. Purification2 Protein purification using Ni-NTA (for 6xHIS-tagged proteins) or glutathione (for GST-tagged proteins) beads at either native or denatured conditions. 1 week Total 4 weeks Note: 1 If customer does not provide the gene, additional $1,000 will be charged (both cloning and sequencing is included) and it may need 1-2 weeks to amplify the gene from library and to confirm the sequences. 2 We do not guarantee the yield of purified protein since it changes from protein to protein. More large-scale culture may be needed to obtain a certain amount of protein for antibody production. Back to top
Protein Expression in Baculoviral System	Download PDF	Order Form
Phase Time I. Cloning of a gene of interest into a baculoviral expression vector1 Amplification/isolation of the gene of interest out of a customer-supplied construct and subcloning it into a baculoviral expression vector. Verification of the authenticity of the subcloned gene by restriction enzyme digest and sequencing. Maxi-prep of the recombinant vector DNA. 2 weeks II. Generation, identification, and plaque purification of baculoviral colonies expressing target proteins Transfection of insect cells (SF9) with the recombinant transfer vector (gene of interest cloned in the vector) and baculoviral DNA. Isolation and plaque-purification of ten baculoviral recombinant clones. Test for expression of the recombinant protein by western blot (customer must provide appropriate antibodies) if desired. 3 weeks III. Amplification of recombinant baculovirus for high titer stock Generation of 500 ml of high-titer viral stock from low-titer stock solution (approximately 10 ml is required) of a single recombinant. A few rounds of amplification of virus may need if the titer of initial virus is too low. Determination of the titer by end-point dilution assay. 1 week IV. Large-scale culture of recombinant baculovirus One liter of insect cells will be infected with high-titer virus stocks (25-30 ml of high titer virus required). 1 week V. Purification2 Protein purification using a Ni-NTA (for 6xHIS-tagged proteins) or glutathione (for GST-tagged proteins) column at either native or denatured condition. 1 week Total 8 weeks Note: 1 If customer does not provide the gene, additional $1,000 will be charged (both cloning and sequencing is included) and it may need 1-2 weeks to amplify the gene from library and to confirm the sequences. 2 We do not guarantee the yield of purified protein since it changes from protein to protein. More large-scale culture may be needed to obtain a certain amount of protein for antibody production. For additional contact tech@abgent.com

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