Abstract
Recombinant Drosophila S2 cells have been used for the expression of many proteins of medical interest. However, membrane-attached glycoproteins, which commonly exhibit lower expression levels compared to soluble proteins, may require special procedures in order to attain high levels of expression. In this study, two S2 cell population enrichment methods (antibiotic and immunomagnetic selection) were evaluated for their ability to enhance expression of the membrane-anchored rabies virus glycoprotein (RVGP). Quantification of RVGP production and determination of its cDNA copy number in transformed cells showed that both enrichment methods increased RVGP expression without significantly affecting its gene copy number. More interestingly, RVGP mRNA levels measured after cycloheximide treatment were poorly correlated with glycoprotein levels. Both enrichment methods enhanced expression of RVGP by recombinant S2 cells, with the highest level of expression achieved using immunomagnetic selection.
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Abbreviations
- RVGP:
-
Rabies virus glycoprotein
- RT-qPCR:
-
Quantitative reverse transcriptase-polymerase chain reaction
- X:
-
Cell concentration (cell/mL)
- S2 RVGP:
-
S2 transfected with RVGP and selected by hygromycin resistence
- S2 RVGP hy:
-
S2 RVGP cultivated for more 28 days with 600 µg/mL of hygromycin
- S2 RVGP imm:
-
S2 RVGP submitted to immunomagnetic selection
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Acknowledgments
This work was financially supported by grants from FAPESP (2012/24647-0), CNPq (402439/2013-9) and Butantan Foundation. C.A. Pereira is recipient of a CNPq 1A senior fellowship.
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Santos, N.G.L., Rocca, M.P., Pereira, C.A. et al. Impact of recombinant Drosophila S2 cell population enrichment on expression of rabies virus glycoprotein. Cytotechnology 68, 2605–2611 (2016). https://doi.org/10.1007/s10616-016-9984-z
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DOI: https://doi.org/10.1007/s10616-016-9984-z