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Establishing elements of a synthetic biology platform for Vaccinia virus production: BioBrickTM design, serum-free virus production and microcarrier-based cultivation of CV-1 cells

Liu, S; Ruban, L; Zhou, Y; Wang, Y; Nesbeth, DN; (2017) Establishing elements of a synthetic biology platform for Vaccinia virus production: BioBrickTM design, serum-free virus production and microcarrier-based cultivation of CV-1 cells. Heliyon , 3 (2) , Article e00238. 10.1016/j.heliyon.2017.e00238. Green open access

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Abstract

Vaccinia virus (VACV) is an established vector for vaccination and is beginning to prove effective as an oncolytic agent. Industrial production of VACV stands to benefit in future from advances made by synthetic biology in genome engineering and standardisation. The CV-1 cell line can be used for VACV propagation and has been used extensively with the CRISPR/Cas9 system for making precise edits of the VACV genome. Here we take first steps toward establishing a scalable synthetic biology platform for VACV production with CV-1 cells featuring standardised biological tools and serum free cell cultivation. We propose a new BioBrick™ plasmid backbone format for inserting transgenes into VACV. We then test the performance of CV-1 cells in propagation of a conventional recombinant Lister strain VACV, VACVL-15 RFP, in a serum-free process. CV-1 cells grown in 5% foetal bovine serum (FBS) Dulbecco’s Modified Eagle Medium (DMEM) were adapted to growth in OptiPRO and VP-SFM brands of serum-free media. Specific growth rates of 0.047 h−1 and 0.044 h−1 were observed for cells adapted to OptiPRO and VP-SFM respectively, compared to 0.035 h−1 in 5% FBS DMEM. Cells adapted to OptiPRO and to 5% FBS DMEM achieved recovery ratios of over 96%, an indication of their robustness to cryopreservation. Cells adapted to VP-SFM showed a recovery ratio of 82%. Virus productivity in static culture, measured as plaque forming units (PFU) per propagator cell, was 75 PFU/cell for cells in 5% FBS DMEM. VP-SFM and OptiPRO adaptation increased VACV production to 150 PFU/cell and 350 PFU/cell respectively. Boosted PFU/cell from OptiPRO-adapted cells persisted when 5% FBS DMEM or OptiPRO medium was observed during the infection step and when titre was measured using cells adapted to 5% FBS DMEM or OptiPRO medium. Finally, OptiPRO-adapted CV-1 cells were successfully cultivated using Cytodex-1 microcarriers to inform future scale up studies.

Type: Article
Title: Establishing elements of a synthetic biology platform for Vaccinia virus production: BioBrickTM design, serum-free virus production and microcarrier-based cultivation of CV-1 cells
Open access status: An open access version is available from UCL Discovery
DOI: 10.1016/j.heliyon.2017.e00238
Publisher version: http://dx.doi.org/10.1016/j.heliyon.2017.e00238
Language: English
Additional information: © 2017 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
Keywords: Biological sciences; Biochemistry; Bioengineering; Cell biology; Microbiology History
UCL classification: UCL
UCL > Provost and Vice Provost Offices > UCL BEAMS
UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Engineering Science
UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Engineering Science > Dept of Biochemical Engineering
URI: https://discovery.ucl.ac.uk/id/eprint/1537823
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