Abstract
The expression enhancement by cytomegalovirus promoter and different intron A (IA) variants were evaluated in CHO-K1, HepG2, HEK-293 and COS-7 cells by assessing the levels of luciferase activity. This data along with mRNA levels measurement indicated that the construct harboring an IA variant with a 200-nucleotide deletion (Δ200) had the greatest impact on increasing luciferase expression among all constructs evaluated. Based on these results, we redesigned pCMV–IA variants and cloned them into plasmids expressing a humanized antibody. These plasmids were then used to transfect CHO-K1 cells. Production of the antibody was not augmented with the Δ200 promoter variant. The 600-nucleotide deletion (Δ600) and whole IA promoter variants expressed similar levels of the recombinant protein. These data indicate that the IA-based enhanced expression of transgenes depends on a small region within the intron.
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Acknowledgments
We thank Izabel CristinaRodrigues Silva for assistance with the statistical analysis. This work was supported by Conselho Nacional do Desenvolvimento Científico e Tecnológico (CNPq) and Financiadora de Estudos e Projetos (FINEP).
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Quilici, L.S., Silva-Pereira, I., Andrade, A.C. et al. A minimal cytomegalovirus intron A variant can improve transgene expression in different mammalian cell lines. Biotechnol Lett 35, 21–27 (2013). https://doi.org/10.1007/s10529-012-1043-z
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DOI: https://doi.org/10.1007/s10529-012-1043-z