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We propose a wide variety of cell lines and expression vectors to optimize your insect cell protein expression.
Unrivalled protein expression expertise
We routinely perform protein expression since 2003. 1500+ proteins expressed so far.
Our baculovirus protein expression service includes optimization of several parameters such as choice of insect cell line, multiplicity of infection and incubation time.
Choose a unique partner from gene synthesis to protein expression.
Benefit from ProteoGenix’s gene synthesis expertise to get an optimized gene for insect cell expression.
PhD account managers
At ProteoGenix, you benefit immediately from the expertise of a PhD account manager!
Expression vector construction (optional)
Virus production and amplification
Small scale protein expression
Protein expression scale up and purification
We were asked by a customer to produce a recombinant Beta-Glucosidase protein using a baculovirus expression system (Insect cell production). Our customer ordered a full Insect cell package, including small-scale expression and purification tests followed by a 1L pilot production and purification. We reached a yield of 370 mg/L and delivered 60 mg after small-scale expression. 1L pilot production was no longer useful and was therefore not charged to our customer. The services carried out included:
Gene synthesis and sequence optimization
Subcloning in BV expression vector
Recombinant Bacmid and virus generation
Recombinant protein expression and purification tests
The cDNA coding for Beta-Glucosidase protein was chemically synthesized after sequence optimization for insect cells expression. It was then subcloned in a proprietary expression vector. A sequence coding for a 6His tag was added for further purification.
Figure 1. Analysis of insect cell expression during P1 generation.
Left. Coomassie Blue Staining. Right. WB with anti-His antibody (ECL substrate) “-“. Un-transfected
control culture. 1 and 2 are transfections with 2 different Bacmid clones
A protein band matching with Beta-Glucosidase protein molecular weight is observed by SDS-PAGE in native protein extracts. This result is confirmed by Western blot (Figure 1, green arrows). Recombinant bacmid clone No1 was used for expression tests.
Figure 2. Protein Expression Tests with P2 stock.
Coomassie Blue Staining. Left. Cell type 1.Right. Cell type 2. “-“. Negative control culture.
Expression tests with P2 confirm that the target is present at high level in native protein extracts (Figure 2, green arrows). Optimal conditions for native expression: Figure 2, red arrow = loi
Figure 3. Protein small-scale purification test. Coomassie Blue Staining.
Left. Purification profile. Right. Final QC of purification pool
IN. Input. FT. Flow through. W1-W3. Washing steps. E1-E9. Eluted fractions.
Recombinant Beta-Glucosidase protein can be produced and purified in native conditions. The purity is ≥ 95%, and the production/purification yield is approximately 370mg/L. Baculovirus expression system was a perfect choice for this protein.
Baculovirus expression systems are eukaryotic expression systems. This confers to these systems all the advantages conferred by their eukaryotic nature such as:
They even include unique advantages compared to other eukaryotic systems such as:
High level of protein expression (even if higher protein yields could be achieved using E. coli or yeast protein expression).
To conclude, insect cells expression represents the best option for researchers and companies willing to produce their protein in a eukaryotic expression system (with glycosylation profiles close to those obtained in mammalian cells) at affordable price.
There are many factors influencing the yield of baculovirus protein expression. One of the most important is the multiplicity of infection (MOI). Multiplicity of infection refers to the number of virus to add to cells.
There are mainly two processes used for baculovirus protein expression:
The yield of insect cell protein expression can also be enhanced by optimizing the incubation time. The goal of this step is to determine at which incubation time the cell culture should be harvested. Basically, cell culture should be harvested before protein degradation starts.
As each project represents a unique challenge, ProteoGenix tests several cell lines, multiplicity of infections, incubation times and culture media (under request). Contact our PhD account manager to get your custom offer.
The baculovirus expression system is based on the infection of insect cells with baculovirus. Baculovirus are insect pathogens controlling the insect population in nature. They present a biphasic replication cycle driven by two forms of the virus:
This latest form of the virus is not necessary for infection of insect cell cultures. Thus, the polh and p10 promoters can be exploited for recombinant protein expression. This is the basis of the baculovirus expression system.
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