flashBAC System compared to other leading systems

flashBAC System
  • Co-transfect flashBAC & transfer vector into Sf9 cells
  • 2 ml P1 stock 1x107-5x107pfu/ml
  • 500 ml P2 stock > 1x108pfu/ml
 
Bac-to-Bac System
  • Transform DNA into EH10Bac E.coli = transposition into bacmid
  • Blue-white selection of recombinant E.coli on antibiotic plates
  • Verify recombinant colonies by PCR
  • Amplify and purify DNA. Transfect µl into 35mm2 dish of Sf9 cells
  • 2ml P1 stock. 2x107 - 4x107 pfu/ml
  • 500 ml P2 stock > 1x108pfu/ml
 
Baculodirect
  • Gene of interest cloned into Gateway entry vector
  • Obtain seed stock
  • Complete plaque assay of seed stock prior to amplification
    to determine titre (optional)
  • Obtain high titre virus stock (day 10 if plaque assay not done)
 
BacPac
  • Co transfect BacPak and transfer vector into cells
  • Seed stock obtained (2ml)
  • Plaque purification completed - 8 days
  • Verify recombinant plaques by PCR. Infect flask for P1 stock
  • Obtain P1 stock
  • Obtain P2 stock (500ml)
 
BacVector 3000 System
  • Co-transfect BacVector 3000 & transfer vector into Sf9 cells
  • 2ml P0 stock. 1x106 - 1x107 pfu/ml
  • 2 rounds plaque purification = 7 days
  • Verify recombinant plaques by PCR. Infect 25 cm2 flask for P1 stock
  • 5ml P1 stock. 1x106 - 1x107 pfu/ml
  • 500 ml P2 stock > 1x108fu/ml
 
Bac-n-Blue System
  • Co-transfect Bac-n-Blue & transfer vector into Sf9 cells
  • 2 ml P1 stock 1x106-1x107pfu/ml
  • 2 rounds plaque purification = 7 days
  • Verify recombinant plaques by PCR.
    Infect 25 cm2 flask for P1 stock
  • Amplify 5ml P1 stock 1x106-1x107pfu/ml
  • 500 ml P2 stock > 1x108pfu/ml
Note: Timescales are taken from manufacturers user manuals
Features flashBAC System Bac-to-Bac System Baculodirect BacPac BacVector 3000 System Bac-n-Blue System
Bacmid Yes Yes Yes Yes    
Contains gene deletions to enhance secretion and improve protein yields Yes       Yes  
Utilises homologous recombination in insect cells so recombinant viruses are stable Yes   Yes Yes Yes Yes
Compatible with standard transfer vectors Yes   Yes Yes Yes  
Doesn’t require plaque purification Yes Yes Yes