Effect of the peak cell density of recombinant AcMNPV-infected Hi5 cells on baculovirus yields

Huynh, Hoai T., Tran, Trinh T. B., Chan, Leslie C. L., Nielsen, Lars K. and Reid, Steven (2014) Effect of the peak cell density of recombinant AcMNPV-infected Hi5 cells on baculovirus yields. Applied Microbiology and Biotechnology, 99 4: 1687-1700. doi:10.1007/s00253-014-6260-z


Author Huynh, Hoai T.
Tran, Trinh T. B.
Chan, Leslie C. L.
Nielsen, Lars K.
Reid, Steven
Title Effect of the peak cell density of recombinant AcMNPV-infected Hi5 cells on baculovirus yields
Journal name Applied Microbiology and Biotechnology   Check publisher's open access policy
ISSN 1432-0614
0175-7598
Publication date 2014-12-05
Year available 2014
Sub-type Article (original research)
DOI 10.1007/s00253-014-6260-z
Open Access Status
Volume 99
Issue 4
Start page 1687
End page 1700
Total pages 14
Place of publication Heidelberg Germany
Publisher Springer
Collection year 2015
Language eng
Formatted abstract
The phenomenon of the cell density effect is not readily explained by an obvious nutrient limitation, and a recent study has suggested that for recombinant Autographa californica multiple nucleopolyhedrovirus (rAcMNPV)-infected Sf9 cells, a drop in messenger RNA (mRNA) levels may be sufficient to explain the cell density effect for this system. The current study aims to investigate the response in cell-specific yields (viral DNA (vDNA), LacZ mRNA and β-galactosidase (β-Gal) protein) with increasing infection cell density (ICD) for rAcMNPV-infected Hi5 cells, where the rAcMNPV expresses the β-Gal gene under control of the polyhedral promoter. Hi5 cells in suspension culture of Express Five® medium were synchronously infected with a rAcMNPV at multiple ICDs between 0.5 and 6 × 106 cells/mL and a multiplicity of infection of 10 plaque-forming units (PFU)/cell either in the original or fresh medium conditions. There were negative correlations between the three key virus infection indicators (vDNA, mRNA and β-Gal) and the peak cell density (PCD). However, unlike infected Sf9 cells, the yield decline started at the lowest PCD investigated (0.6 × 106 cells/mL). Generally, the yield decline with increasing PCD was most pronounced for β-Gal followed by mRNA and was more moderate for vDNA. The decline was significantly reduced but not totally arrested when fresh medium replacement was used. The results suggest that the reduction in recombinant protein-specific yields at high PCDs is associated with limitations during the up-stream processes of replication and transcription rather than entirely caused by limitations during translation. In addition, low production rates at late infection stages of moderate to high ICDs are a probable cause of the cell density effect.
Keyword Baculovirus
Hi5 cells
Cell density effect
Viral DNA replication
mRNA expression
β-Gal
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ
Additional Notes Published online ahead of print 05 Dec 2014

Document type: Journal Article
Sub-type: Article (original research)
Collections: Official 2015 Collection
Australian Institute for Bioengineering and Nanotechnology Publications
 
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