Screening of Domestic Silkworm Strains for Efficient Heterologous Protein Expression by Bombyx mori Nuclear Polyhedrosis Virus (BmNPV)
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Recombinant proteins can be generated quickly and easily in large amounts and at low-costin silkworm larvae by using Bombyx mori nuclear polyhedrosis virus (BmNPV). We searchedfor high-permissive silkworm strains that have high production levels of heterologous proteinsand are thus suitable for use as biofactories. In this study, we performed the analysis usinga BmNPV vector expressing luciferase as a marker, and we confirmed protein expressionby evaluating luciferase activity, determined by western blotting and luciferase ELISA, andconfirmed transcription expression by semi- and quantitative real time PCR. For the selectionof host silkworm strains, we first chose 52 domestic BmNPV sensitive strains and thenidentified 10 high-permissive and 5 low-permissive strains. In addition, to determine whichhybrid of the high-permissive strains would show heterosis, nine strains derived through threewaycrossing were tested for luciferase activity by western blotting, and luciferase ELISA. We found a correlation between luciferase activity and luciferase protein expression, but nottranscription. There was no noticeable difference in protein expression levels between Jam313as the high-permissive control strain and the three-way hybrid strains; however, the three-waycross strains showed lower luciferase activity compared with Jam313. In this study, luciferaseprotein production in the larvae of 52 domestic silkworm strains was elucidated using BmNPV.
Abstract
Recombinant proteins can be generated quickly and easily in large amounts and at low-costin silkworm larvae by using Bombyx mori nuclear polyhedrosis virus (BmNPV). We searchedfor high-permissive silkworm strains that have high production levels of heterologous proteinsand are thus suitable for use as biofactories. In this study, we performed the analysis usinga BmNPV vector expressing luciferase as a marker, and we confirmed protein expressionby evaluating luciferase activity, determined by western blotting and luciferase ELISA, andconfirmed transcription expression by semi- and quantitative real time PCR. For the selectionof host silkworm strains, we first chose 52 domestic BmNPV sensitive strains and thenidentified 10 high-permissive and 5 low-permissive strains. In addition, to determine whichhybrid of the high-permissive strains would show heterosis, nine strains derived through threewaycrossing were tested for luciferase activity by western blotting, and luciferase ELISA. We found a correlation between luciferase activity and luciferase protein expression, but nottranscription. There was no noticeable difference in protein expression levels between Jam313as the high-permissive control strain and the three-way hybrid strains; however, the three-waycross strains showed lower luciferase activity compared with Jam313. In this study, luciferaseprotein production in the larvae of 52 domestic silkworm strains was elucidated using BmNPV.
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