Abstract
Bombyx mori ovary-derived BmN4 cells have been successfully adapted to a commercial serum-free medium (SFM; SF900-II) by gradually reducing the serum-containing TC-100 medium content from 100 to 0% (v/v). The BmN4 cells adapted to the SFM (BmN-SFM) adhered strongly to the culture flask and showed altered cell morphology. The BmN-SFM was subcultured 200 times, and the population doubling time was 4.70 d. Infection studies showed that BmN-SFM cells were easily susceptible to B. mori nucleopolyhedrovirus (BmNPV), and both the multiplication of budded virus and the promoter activity of the polyhedrin gene in BmN-SFM cells were almost the same as those in BmN4 cells before adaptation. Additionally, mouse interleukin-3 expressed by a recombinant BmNPV was normally secreted and modified with N-linked glycans in BmN-SFM cells. These findings indicate that BmN-SFM is particularly useful for a BmNPV-based baculovirus expression vector system with serum-free conditions.
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Acknowledgment
This work was supported in part by A-step feasibility study program from Japan Science and Technology Agency (JST) (to MI; 241FT0093).
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Editor: T. Okamoto
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Supplementary figure 1
RAPD profiles of five different cell lines using three primers: GEN1, GEN5, and GEN7. The number of cell lines is as follows: 1, BmN4 (B. mori, ovary); 2, BmN-SFM (B. mori, ovary); 3, Sf-9 (S. frugiperda, ovary); 4, NIAS-BoMa-529b (B. mandarina, fat body); 5, BmVF (B. mori, embryo). (PPTX 187 kb)
Supplementary figure 2
Growth curve of the B. mori cell lines. BmN4 (open columns) and BmN-SFM (closed columns) were seeded on culture dishes, and the cell numbers were evaluated at 24-h intervals. The results represent the average of three independent experiments; standard errors are indicated. (PPTX 58 kb)
Supplementary figure 3
Susceptibility of BmN-SFM cells to BmNPV. The morphology of BmN4 and BmN-SFM cells at 96 hours post-BmNPV-infection. The scale bar is 50 μm. (PPTX 820 kb)
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Iwanaga, M., Adachi, Y., Uchiyama, K. et al. Long-term adaptation of the Bombyx mori BmN4 cell line to grow in serum-free culture. In Vitro Cell.Dev.Biol.-Animal 50, 792–796 (2014). https://doi.org/10.1007/s11626-014-9781-y
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DOI: https://doi.org/10.1007/s11626-014-9781-y