Insect Baculovirus Expression System

Introduction:

Insect baculovirus expression vector system (BEVS) belongs to the eukaryotic expression system, and it’s an expression system with high safety. It has a large genome, which enables the insertion of large exogenous genes, therefore has the great advantage of expressing proteins with large molecular weight. It also has the ability to achieve complete post-translational modification and efficiently express exogenous genes. The system consists of transfer vector, baculovirus vector and the host cell. The system uses one or more baculovirus super-strong promoters, and gets the recombinant virus after the exogenous target gene is inserted into the promoter. The highly efficient expression of the exogenous gene is achieved while the recombinant viruses replicate themselves in the insect cells. BEVS is widely used in virus vaccine development (such as the development of influenza virus vaccine and HPV vaccine), preparation of cell signaling proteins and cytokines, as well as kinase development, etc.

Advantages:
  • Large capacity: ability to carry large gene fragment; advantage in large protein expression
  • High safety: baculovirus has strict species specificity
  • High expression efficiency: the protein can be efficiently expressed in the late-stage infected cells
  • The post-translational modification of the expressed product is similar to that of mammalian cells, particularly the glycosylation; the protein is more likely to be bioactive
  • Baculovirus is easier to amplify and can produce recombinant proteins in large scale
  • Our unique bacmid based expression system has prominent features including low cost, large volume suspension transfection, high titer, shorten lead-time by 1-2 weeks compare to classic process cycle
  • As low as $980, delivery time as short as 7 weeks, yield up to 100 mg/L
Guarantee:

Risk-free: We do NOT charge if we cannot deliver the protein.

Service Process:

Steps Project Process Cusabio Features Lead Time
1 Plasmid construction Codon optimization; gene synthesis Vector optimization
In order to improve the success rate of expression and achieve higher yield, we provide protein expression using C-terminal fusion tags in addition to conventional N-terminal tags, which retains the bioactivity of the protein while ensuring high purity.
15-20 business days
The PCR product is ligated to the expression vectors e.g. pFastbac1-KHM, pFastBac1-MBP, etc.
Transform TOP10 E.coil competent cells
Obtain the correct recombinant plasmid
2 Preparation of recombinant Bacmid and high titer virus Transform DH10Bac cells to get recombinant Bacmid; PCR analysis; Isolate recombinant bacmid DNA Suspension transfection
Unique suspension transfection method greatly increases the protein expression level, and effectively shortens the experimental cycle.
12-15 business days
Transfect recombinant Bacmid DNA into insect cells to obtain baculovirus, and detect expression level by SDS-PAGE; Repeat the infection if necessary
3 Scale up expression and purification Infect insect cells with appropriate baculovirus Expression optimization
After optimization, the large amount of protein can be obtained by infecting host cells with low-passage virus.
5-10 business days
The target protein is purified by affinity chromatography, ion exchange, hydrophobic and molecular sieves.
4 Additional services (optional) Charge Tag removal by restriction digestion Flexible additional services
Customers can flexibly choose from a variety of additional services to their specific needs, e.g. Endotoxin removal, Filter-sterilization, Tag removal, Lyophilization, etc. Some are complimentary, and some require additional charge.
3 business days
Free Filter-sterilization; Endotoxin removal; Lyophilization (Note: Lyophilization and Filter-sterilization can’t be met simultaneously) 2 business days
5 Quality Control Testing of purity, concentration, etc. QC report is provided. Detailed COA report
Detailed product data sheet and COA are provided for each project.
3-5 business days
Total lead time 35-50 business days

Case 1
The protein was highly expressed in our company's Insect baculovirus expression vector system. A clear band was observed from cell lysate by SDS-PAGE. The yield was up to 20 mg/L after purification.

  • Protein purification image
  • Lane 1: Flow through.
  • Lane 2: Cell lysate
  • Lane 3: Marker
  • Lane 4: 30 mM imidazole elution
  • Lane 5: 60 mM imidazole elution
  • Lane 6: 250 mM imidazole elution

Case 2
This target protein is relatively large. After gene synthesis, vector construction, bacmid construction, we finally produced the protein in sf9 cells. The yield reached to 5 mg/L, and the purity was 95%.

  • Protein purification image
  • Lane 1:60 mM imidazole elution

Case 3
The molecular weight of this target protein is relatively small, and it is quite difficult to express. We chose to use the pFastBac1-MBP vector to make the recombinant construct. After enzyme digestion and secondary purification, the purity of target protein reached 95%.

  • Protein purification image
  • Lane 1: Fusion protein before digestion
  • Lane 2: Fusion protein after digestion
  • Lane 3: Fusion protein after secondary purification

Characteristic expression systems

pFastbac1-KHM vector + sf9 cells, highly efficient expression system

  • 1.

    Seamless cloning, no restriction enzyme needed.

  • 2.

    Compared with conventional 6xHis tag, the N-terminal 10xHis tag has a stronger binding ability in IMAC. The thrombin site makes it easy to remove the tag.

  • 3.

    The C-terminal MYC-tag can be used for WB detection, and it has stronger sensitivity compared to His-tag.

  • 4.

    Amp resistance screening.

pFastBac1-MBP vector+sf9 cell, highly efficient expression system

  • 1.

    This vector contains MBP fusion tag that has a strong ability to promote expression.

  • 2.

    It is a better system for small protein expression.

  • 3.

    Seamless cloning, no restriction enzyme needed.

  • 4.

    The TEV cleavage site makes it easy for tag removal.

  • 5.

    Amp resistance screening.

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