Efficient large-scale protein production of larvae and pupae of silkworm by Bombyx mori nuclear polyhedrosis virus bacmid system

Motohashi, Tomoko; Shimojima, Tsukasa; Fukagawa, Tatsuo; Maenaka, Katsumi; Park, Enoch Y.

doi:10.1016/j.bbrc.2004.11.060

Cited by 195 publications

(169 citation statements)

References 8 publications

Supporting

Mentioning

163

Contrasting

Unclassified

Order By: Relevance

“…In order to save the labor required to generate the recombinant Bombyx mori nuclear polyhedrosis viruses (BmNPVs), we used the Gateway cloning system (Invitrogen) and the Bac-to-Bac system [13]. For the first step, six entry plasmids with distinct signal peptide coding sequences were constructed (Figure 1).…”

Section: Signal Peptides Used For Secretion In the Baculovirus-silkwomentioning

confidence: 99%

See 1 more Smart Citation

Comparison of signal peptides for efficient protein secretion in the baculovirus-silkworm system

et al. 2013

View full text Add to dashboard Cite

The baculovirus-silkworm expression system is widely used as a mass production system for recombinant secretory proteins. However, the final yields of some recombinant proteins are not sufficient for industrial use. In this study, we focused on the signal peptide as a key factor for improving the efficiency of protein production. Endoplasmic reticulum (ER) translocation of newly synthesized proteins is the first stage of the secretion pathway; therefore, the selection of an efficient signal peptide would lead to the efficient secretion of recombinant proteins. The Drosophila Bip and honeybee melittin signal peptides have often been used in this system, but to the best of our knowledge, there has been no study comparing secretion efficiency between exogenous and endogenous signal peptides. In this study we employed signal peptides from 30K Da and SP2 proteins as endogenous signals, and compared secretion efficiency with those of exogenous or synthetic origins. We have found that the endogenous secretory signal from the 30K Da protein is the most efficient for recombinant secretory protein production in the baculovirus-silkworm expression system.

show abstract

Section: Signal Peptides Used For Secretion In the Baculovirus-silkwomentioning

confidence: 99%

“…The pDEST8 based transfer vectors obtained were transformed into E. coli BmDH10Bac cells [13]. Recombinant bacmid DNAs were purified using the FlexiPrep kit (GE Healthcare Bioscience, Piscataway, USA) and transfected into Bme21 cells (9×10 5 cells per well) [14] with the CellFECTIN reagent (Invitrogen).…”

Section: Construction Of Recombinant Baculovirusmentioning

confidence: 99%

Comparison of signal peptides for efficient protein secretion in the baculovirus-silkworm system

et al. 2013

View full text Add to dashboard Cite

show abstract

“…Two decades ago, protein expression in silkworms was difficult and time-consuming, because of cultured insect cells were mainly used as baculovirus hosts for protein production. However, Bombyx mori nucleopolyhedrovirus (BmNPV) bacmid (a baculovirus shuttle vector), which can be replicated in E. coli and generate the recombinant baculovirus DNA by site-specific transposition in E.coli, has been developed [68] and many reports on recombinant protein expression in silkworms are being published [69].…”

Section: Vlps Production Using Silkworm Larvae and Their Applicationsmentioning

confidence: 99%

Functional Virus-Like Particles Production Using Silkworm and Their Application in Life Science

Kato¹,

Deo²,

Park

et al. 2012

J Biotechnol Biomaterial

View full text Add to dashboard Cite

Virus-like particles (VLPs), which mimic the structure of authentic virus, are of significant importance owing to their wide ranging applications. VLPs have the potential to serve as candidate vaccine in addition to subunit vaccines and also serve as a nano-bioparticle scaffold for displaying complex foreign proteins. Here, we review different methods available to produce VLPs with various host expression systems including from microorganisms to mammalian cells and the current potential of silkworms as producers of these VLPs. Recently, silkworms have been used for efficient production of VLPs too in addition to mass production of recombinant proteins, which have contributed to molecular structural analysis, molecule-molecule interactions in biology and biotechnology.

show abstract

“…Recently, Bac-to-Bac/BmNPV baculovirus expression system has been developed (Motohashi et al 2005). In this system, recombinant bacmid baculoviruses were generated by site-specific transposition in E. coli which needs no more than 10 days due to the elimination of multiple rounds of purification of viruses, and the Bm-NPV bacmid system can be applied in Bm cell lines and silkworm.…”

Section: Introductionmentioning

confidence: 99%

Expression of EGFP driven by BmNPV orf4 promoter, a novel immediate early promoter

Shen

et al. 2009

Biologia

View full text Add to dashboard Cite

Bombyx mori nucleopolyhedrovirus (BmNPV) orf4 has been shown to be expressed at very early stage of Bm-NPV infection cycle. In this study, using transient expression experiment, we demonstrated for the first time that orf4 promoter is an immediate early promoter, indicating that orf4 may play a role in the immediate-early stage of BmNPV infection. Moreover, with the recently developed Bac-to-Bac/BmNPV baculovirus expression system and a modified pFastBac1 whose polyhedrin promoter was replaced with orf4 promoter, a recombinant bacmid baculovirus expressing enhanced green fluorescent protein (EGFP) under the control of orf4 promoter in Bombyx mori (Bm) cells was successfully constructed. The result not only showed that the polyhedrin promoter can be replaced easily with other promoters to direct the expression of foreign genes by using this novel system but also laid the foundation for the rescue experiment of orf4 deletion mutant.

show abstract

Efficient large-scale protein production of larvae and pupae of silkworm by Bombyx mori nuclear polyhedrosis virus bacmid system

Cited by 195 publications

References 8 publications

Comparison of signal peptides for efficient protein secretion in the baculovirus-silkworm system

Comparison of signal peptides for efficient protein secretion in the baculovirus-silkworm system

Functional Virus-Like Particles Production Using Silkworm and Their Application in Life Science

Expression of EGFP driven by BmNPV orf4 promoter, a novel immediate early promoter

Contact Info

Product

Resources

About