Human single-chain antibody expression in the hemolymph and fat body of silkworm larvae and pupae using BmNPV bacmids
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概要
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Seven sets of recombinant expression vectors were constructed for the expression of the human anti-bovine serum albumin (BSA) single-chain Fv fragment (scFv) 13CG2 fused to the C terminus of GFP_<uv> or bombyxin (bx) signal peptide in the hemolymph and fat body of silkworm larvae and pupae, using cysteine protease- (BmNPV-CP^-), and cysteine protease- and chitinase-deficient (BmNPV-CP^-Chi^-) bacmids. When BmNPV-CP^- or BmNPV-CP^-Chi^- bacmids were used, 16.9-18.9mg/l (11.6-15.0μg/larva) of scFv was expressed at 6d.p.i., whereas wild-type BmNPV bacmid expressed only 4.4mg/l, probably because of proteolytic degradation of the protein. The scFv yield in silkworm pupae was only 0.67-1.0μg/pupa, which was 5.4% of that in the hemolymph of silkworm larvae. The bx signal peptide enabled the secretion of scFv into the hemolymph. Without the signal sequence, the fusion protein accumulated in the fat body and lost its biological function. The removal of GFP_<uv> significantly increased the scFv yield in the hemolymph of silkworms to 188.4mg/l (132.4mg/larva), which was ten times higher than that of the fusion protein.
著者
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UEDA Hiroshi
Department of Pathology, School of Medicine Gifu University
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Park Enoch
Laboratory Of Biotechnology Department Of Applied Biological Chemistry Shizuoka University:integrate
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Kato Tatsuya
Laboratory of Biotechnology, Faculty of Agriculture, Shizuoka University
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Park Enoch
Laboratory Of Biotechnology Department Of Applied Biological Chemistry Faculty Of Agriculture Shizuo
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Kato Tatsuya
Shizuoka Univ. Shizuoka Jpn
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Nishina Takuya
Laboratory Of Biotechnology Department Of Applied Biological Chemistry Shizuoka University
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Ishikiriyama Motoki
Laboratory of Biotechnology, Department of Applied Biological Chemistry, Shizuoka University
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Kato Tatsuya
Laboratory Of Biotechnology Department Of Applied Biological Chemistry Shizuoka University
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Ishikiriyama Motoki
Laboratory Of Biotechnology Department Of Applied Biological Chemistry Shizuoka University
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Ueda Hiroshi
Department Of Dermatology Fujita Health University School Of Medicine
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Kato Tatsuya
Laboratory Of Biotechnology Department Of Applied Biological Chemistry Faculty Of Agriculture Shizuo
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Ueda Hiroshi
Department Of Chemistry And Biotechnology School Of Engineering University Of Tokyo
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Ueda Hiroshi
Department Of Anatomy University Of Occupational And Environmental Health School Of Medicine
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Ueda Hiroshi
Department Of Anatomy School Of Medicine University Of Occupational And Environmental Health
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