Skip to main content
SpringerLink
Log in

Establishment of a Bombyx mori nucleopolyhedrovirus (BmNPV) hyper-sensitive cell line from the silkworm e21 strain

  • Original Research Paper
  • Published:
Biotechnology Letters Aims and scope Submit manuscript

Abstract

Baculoviral expression systems, including those of Autographa californica multiple nucleopolyhedrovirus Bombyx mori nucleopolyhedrovirus (BmNPV), are used for recombinant protein production. Four B. mori-derived (BmN4, Bm5, Bmc140, and Bme21) cell lines were infected with recombinant BmNPV viruses expressing firefly luciferase or EGFP as reporters under the control of a viral polyhedrin promoter. Bme21 exhibited significantly higher (100-fold) luciferase activity than BmN4 and Bm5. With the EGFP reporter protein, Bme21 cells showed a marked increase in the ratio of EGFP-positive cells, reaching 90 % on day 4 post-infection, while Bm5 and BmN4 cells had a slow increase in the ratio of their EGFP-positive population. The viral titer in a supernatant of Bme21 cell culture increased faster than those of Bm5 and BmN4 cells. This susceptibility indicates that the Bme21 cell line is useful for large-scale protein expression using BmNPV.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3

Similar content being viewed by others

References

  • Airenne KJ, Makkonen KE, Mähönen AJ, Ylä-Herttuala S (2011) Baculoviruses mediate efficient gene expression in a wide range of vertebrate cells. Methods Mol Biol 737:279–301

    Article  PubMed  CAS  Google Scholar 

  • Bonning BC, Hammock BD (1992) Development and potential of genetically engineered viral insecticides. Biotechnol Genet Eng Rev 10:455–489

    PubMed  CAS  Google Scholar 

  • Chen TH, Lo YP, Yang CF, Chen WJ (2012) Additive protection by antioxidant and apoptosis-inhibiting effects on mosquito cells with dengue 2 virus infection. PLoS Negl Trop Dis 6:e1613

    Article  PubMed  CAS  Google Scholar 

  • Granados RR, Li GX, Derksen ACG, McKenna KA (1994) A new insect-cell line from Trichoplusia ni (BTI-TN-5B1-4) susceptible to Trichoplusia ni single enveloped nuclear polyhedrosis-virus. J Invertebr Pathol 64:260–266

    Article  Google Scholar 

  • Hitchman RB, Murguía-Meca F, Locanto E, Danquah J, King LA (2011) Baculovirus as vectors for human cells and applications in organ transplantation. J Invertebr Pathol 107:S49–S58

    Article  PubMed  CAS  Google Scholar 

  • Imanishi S, Kobayashi J, Sekine T (2012) Serum-free culture of an embryonic cell line from Bombyx mori and reinforcement of susceptibility of a recombinant BmNPV by cooling. In Vitro Cell Dev Biol Anim 48:137–142

    Article  PubMed  CAS  Google Scholar 

  • Ishikawa H, Ikeda M, Alves CA, Thiem SM, Kobayashi M (2004) Host range factor 1 from Lymantria dispar nucleopolyhedrovirus (NPV) is an essential viral factor required for productive infection of NPVs in IPLB-Ld652Y cells derived from L. dispar. J Virol 78:12703–12708

    Article  PubMed  CAS  Google Scholar 

  • Kajikawa M, Sasaki K, Wakimoto Y, Toyooka M, Motohashi T, Shimojima T, Takeda S, Park EY, Maenaka K (2009) Efficient silkworm expression of human GPCR (nociceptin receptor) by a Bombyx mori bacmid DNA system. Biochem Biophys Res Commun 385:375–379

    Article  PubMed  CAS  Google Scholar 

  • Khurad AM, Kanginakudru S, Qureshi SO, Rathod MK, Rai MM, Nagaraju J (2006) A new Bombyx mori larval ovarian cell line highly susceptible to nucleopolyhedrovirus. J Invertebr Pathol 92:59–65

    Article  PubMed  CAS  Google Scholar 

  • Khurad AM, Zhang MJ, Deshmukh CG, Bahekar RS, Tiple AD, Zhang CX (2009) A new continuous cell line from larval ovaries of silkworm, Bombyx mori. In Vitro Cell Dev Biol Anim 45:414–419

    Article  PubMed  Google Scholar 

  • Maeda S (1989) Expression of foreign genes in insects using baculovirus vectors. Annu Rev Entomol 34:351–372

    Article  PubMed  CAS  Google Scholar 

  • Miao Y, Zhang Y, Nakagaki K, Zhao T, Zhao A, Meng Y, Nakagaki M, Park EY, Maenaka K (2006) Expression of spider flagelliform silk protein in Bombyx mori cell line by a novel Bac-to-Bac/BmNPV baculovirus expression system. Appl Microbiol Biotechnol 71:192–199

    Article  PubMed  CAS  Google Scholar 

  • Motohashi T, Shimojima T, Fukagawa T, Maenaka K, Park EY (2005) Efficient large-scale protein production of larvae and pupae of silkworm by Bombyx mori nuclear polyhedrosis virus bacmid system. Biochem Biophys Res Commun 326:564–569

    Article  PubMed  CAS  Google Scholar 

  • O’Reilly DR, Miller LK, Luckow VA (eds) (1992) Baculovirus expression vectors. A laboratory manual. WH Freeman, New York

    Google Scholar 

  • Pan MH, Xiao SQ, Chen M, Hong XJ, Lu C (2007) Establishment and characterization of two embryonic cell lines of Bombyx mori. In Vitro Cell Dev Biol Anim 43:101–104

    Article  PubMed  CAS  Google Scholar 

  • Pandharipande TN (1994) Two new continuously growing cell lines from Bombyx mori (Lepidoptera: Bombycidae). Appl Entomol Zool 29:604–607

    Google Scholar 

  • Schwartz PS, Chen CS, Waxman DJ (2002) Enhanced bystander cytotoxicity of P450 gene-directed enzyme prodrug therapy by expression of the antiapoptotic factor p35. Cancer Res 62:6928–6937

    PubMed  CAS  Google Scholar 

  • Sudeep AB, Mishra AC, Shouche YS, Pant U, Mourya DT (2002) Establishment of two new cell lines from Bombyx mori (L.) (Lepidoptera: Bombycidae) and their susceptibility to baculoviruses. Indian J Med Res 115:189–193

    PubMed  CAS  Google Scholar 

  • Vaughn JL, Goodwin RH, Tompkins GJ, McCawley P (1977) The establishment of two cell lines from the insect Spodoptera frugiperda (Lepidoptera: Noctuidae). In Vitro Cell Dev Biol Plant 13:213–217

    Article  CAS  Google Scholar 

  • Wang C, Smith RL (1975) Lowry determination of protein in the presence of Triton X-100. Anal Biochem 63:414–417

    Article  PubMed  CAS  Google Scholar 

  • Wilson JA, Forney SD, Ricci AM, Allen EG, Hefferon KL, Miller LK (2005) Expression and mutational analysis of Autographa californica nucleopolyhedrovirus HCF-1: functional requirements for cysteine residues. J Virol 79:13900–13914

    Article  PubMed  CAS  Google Scholar 

  • Yu M, Carstens EB (2012) Choristoneura fumiferana multiple nucleopolyhedrovirus LEF-3-P143 complex can complement DNA replication and budded virus in an AcMNPV LEF-3-P143 double knockout bacmid. J Gen Virol 93:383–388

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

This work was supported in part by KAKENHI Nos. 22248003, 22248004 and 23580077 from the Japan Society for the Promotion of Science. The cost of publication was supported in part by the Research Grant for Young Investigators of Faculty of Agriculture, Kyushu University. Cultured silkworm cell lines used in this study were kindly provided by Dr. Aoki, Kyushu University.

Author information

Authors and Affiliations

  1. Laboratory of Silkworm Science, Kyushu University Graduate School of Bioresource and Bioenvironmental Sciences, 6-10-1 Hakozaki, Higashi-ku, Fukuoka, 812-8581, Japan

    Jae Man Lee, Naoya Kawakami, Hiroaki Mon, Hitoshi Mitsunobu & Takahiro Kusakabe

  2. Institute of Biological Control, Kyushu University Graduate School of Bioresource and Bioenvironmental Sciences, 6-10-1 Hakozaki, Higashi-ku, Fukuoka, 812-8581, Japan

    Kazuhiro Iiyama

  3. Department of Biological Production, Tokyo University of Agriculture and Technology, Fuchu, Tokyo, 183-8538, Japan

    Satoshi Ninaki

  4. Division of Structural Biology, Medical Institute of Bioregulation, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka, 812-8582, Japan

    Katsumi Maenaka

  5. Department of Applied Biological Chemistry, Faculty of Agriculture, Shizuoka University, 836 Ohya, Shizuoka, 422-8529, Japan

    Enoch Y. Park

Authors
  1. Jae Man Lee
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Naoya Kawakami
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Hiroaki Mon
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Hitoshi Mitsunobu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Kazuhiro Iiyama
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Satoshi Ninaki
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Katsumi Maenaka
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Enoch Y. Park
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Takahiro Kusakabe
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Takahiro Kusakabe.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (DOCX 170 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Lee, J.M., Kawakami, N., Mon, H. et al. Establishment of a Bombyx mori nucleopolyhedrovirus (BmNPV) hyper-sensitive cell line from the silkworm e21 strain. Biotechnol Lett 34, 1773–1779 (2012). https://doi.org/10.1007/s10529-012-0971-y

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10529-012-0971-y

Keywords

Search

Navigation