Insect System
Baculovirus expression systems have several known advantages over E. coli system, such as improved solubility, ability to incorporate post-translational modifications, and higher yields for secreted proteins. For customers who prefer a eukaryotic expression system, are exploring lower-cost alternatives to mammalian expression system but do not want to compromise on the overall quality of their recombinant protein, we strongly recommend considering our baculovirus expression system. Using our baculovirus expression system we can establish the best possible protocol for your custom project and then scale up to get the highest expression of your target protein.

Phase 1: Subcloning into Recombinant Baculovirus Transfer Vector

Phase 2: Generation and Evaluation of Baculovirus Recombinants

Phase 3: Amplification of Recombinant Baculovirus Stock

Phase 4: Baculovirus Expression Optimization Service

Phase 5: Protein Production Service

Phase 6: Protein Extraction & Purification

Phase 1 Subcloning into Recombinant Baculovirus Transfer Vector

Description:

  • Design/construct the gene of interest into the elected recombinant Baculovirus transfer vector
  • Verify the subcloned gene by restriction enzyme digest and sequencing
  • Prepare the recombinant vector DNA suitable for co-transfection with our proprietary baculovirus DNA

Required Materials:

  • 5 ug of plasmid containing gene of interest plus complete vector sequence in electronic format
  • Target protein/cDNA sequence (codon optimization and gene synthesis)

Deliverables:

  • 5 ug of ready-to-use recombinant Baculovirus transfer plasmid containing the gene of interest
  • Verified sequence of baculovirus transfer plasmid containing the gene of interest

Time Frame:

  • 2 weeks：If insert > 2kb, one week additional per 1kb

Price Estimate:

  • $960: If insert > 2kb, $350 additional per 1kb

Phase 2 Generation and Evaluation of Baculovirus Recombinants

Description:

  • Cotransfect recombinant Baculovirus transfer vector containing gene of interest with 3 different proprietary linearized Baculovirus DNAs using BaculoShuttle transfection reagent
  • Identify and amplify three recombinant constructs
  • Select the best recombinant baculovirus construct for the protein expression
  • Amplify three rounds to generate a high titer virus stock (>1e8 pfu/ml)
  • Titrate high titer stock using end point dilution

Required Materials:

  • 10 ug of ready to use recombinant Baculovirus transfer plasmid containing the gene of interest

Deliverables:

  • Three cell pellets or baculovirus-conditioned insect cell media for evaluating Baculovirus-mediated protein expression
  • 30ml of ready-to-use baculovirus high titer stock (>1e8 opfu/ml)

Time Frame:

  • 2-4 Weeks

Price Estimate:

  • $1,750

Phase 3 Amplification of Recombinant Baculovirus Stock

Description:

  • Amplify low titer recombinant Baculovirus to high titer stock (>1e8 pfu/ml)
  • Titrate high-titer stock by end point dilution

Required Materials:

  • 5 ml of high-titer (>1e8 pfu) recombinant Baculovirus stock

Deliverables:

  • 500 ml of high-titer recombinant Baculovirus stock solution

Time Frame:

  • 2 - 3 Weeks

Price Estimate:

  • $540 per 500ml

Phase 4 Baculovirus Expression Optimization Service

Description:

  • Prepare/grow log phase insect cultures
  • Grow adherent insect cultures on Phoenix Tissue Culture dishes
  • Infect three insect cell lines (Sf21, Sf9, and T.ni)
  • Infect at low and high multiplicity of infection (MOI)
  • Harvest cells and Baculovirus-conditioned cell media at 48hr and 72hr post-infection

Required Materials:

  • At least 5-10 ml of recombinant Baculovirus stock

Deliverables:

  • 18 pairs of cell pellets and virus conditioned cell media for evaluation of protein expression

Time Frame:

  • 2 - 4 Days

Price Estimate:

  • $1,800

Phase 5 Protein Production Service

Description:

  • Prepare high titer recombinant Baculovirus stock for protein production
  • Prepare adherent cell culture using Phoenix Tissue Culture dishes (~34 large 150x25mm dish/liter)
  • Infect cells under select conditions
  • Harvest/process cells or conditioned media at selected time postinfection

Required Materials:

  • High titer recombinant Baculovirus (25 ml per liter of protein production desired)

Deliverables:

  • Cell pellets or conditioned cell media containing target protein

Time Frame:

  • 2~3 weeks (1~5 Liters)
  • 4~5 weeks (5~20 Liters)
  • 8~10 weeks (20 Liters and above)

Price Estimate:

  • $560/Liter
  • For multiple liters please ask for quotation

Phase 6 Protein Extraction & Purification

Description:

  • Process cell pellet or cell media from 1 liter of insect using our proprietary reagents and SOP
  • Purify either under native or denatured conditions using appropriate affinity resin for 6xHis or GST-tagged recombinant proteins
  • SDS PAGE analysis
  • Dialyze/buffer exchange purified protein in storage buffer

Required Materials:

  • Cell pellet or baculovirus-conditioned cell media containing the target protein

Deliverables:

  • Purified protein in standard or customer-specified storage buffer
  • SDS-PAGE data

Time Frame:

  • 2 - 3 Weeks

Price Estimate:

  • $1,350 for 1 Liter
  • Inquire for larger quantities