Versatility of chitosan/BmNPV bacmid DNA nanocomplex as transfection reagent of recombinant protein expression in silkworm larvae.
OBJECTIVE
To examine the feasibility of chitosan as an alternative transfection reagent candidate for protein expression in Bm5 cells and silkworm larvae using recombinant BmNPV bacmid DNA.


RESULTS
Chitosan 100 and recombinant Bombyx mori nucleopolyhedrovirus (BmNPV) bacmid DNA, in amino group/phosphate group (N/P) ratios of 0.1-10, were used for formation of chitosan/DNA nanocomplexes. The chitosan/BmNPV bacmid DNA nanocomplexes showed higher specific activity of GFPuv-β1,3-N-acetylglucosaminyltransferase 2 (β3GnT2) fusion protein (GGT2) expressed in silkworm larvae than DMRIE-C, a conventional silkworm transfection reagent. In particular, the composition of chitosan and BmNPV bacmid DNA nanocomplexes formed by an N/P ratio of 8 or 10, respectively, showed the highest specific activity of β3GnT2 in the silkworm larvae hemolymph. In addition, three different proteins were expressed in silkworm larvae to the same extent using chitosan as that using DMRIE-C.


CONCLUSIONS
This is the first finding that chitosan/BmNPV bacmid DNA nanocomplexes can rival the performance of commercially available transfection reagents for the expression of recombinant proteins in Bm5 cells and silkworm larvae.
Laboratory of Biotechnology, Green Chemistry Research Division, Research Institute of Green Science and Technology, Shizuoka University, 836 Ohya Suruga-ku, Shizuoka, 422-8529, Japan.|Laboratory of Biotechnology, Department of Bioscience, Graduate School of Science and Technology, Shizuoka University, 836 Ohya, Suruga-ku, Shizuoka, 422-8529, Japan.|Laboratory of Biotechnology, Department of Applied Biological Chemistry, Faculty of Agriculture, Shizuoka University, 836 Ohya, Suruga-ku, Shizuoka, 422-8529, Japan.
Tatsuya
T
Kato
Laboratory of Biotechnology, Department of Applied Biological Chemistry, Faculty of Agriculture, Shizuoka University, 836 Ohya, Suruga-ku, Shizuoka, 422-8529, Japan.
Sho
S
Arai
Laboratory of Biotechnology, Department of Applied Biological Chemistry, Faculty of Agriculture, Shizuoka University, 836 Ohya, Suruga-ku, Shizuoka, 422-8529, Japan.
Hirono
H
Ichikawa
Laboratory of Biotechnology, Green Chemistry Research Division, Research Institute of Green Science and Technology, Shizuoka University, 836 Ohya Suruga-ku, Shizuoka, 422-8529, Japan. park.enoch@shizuoka.ac.jp.|Laboratory of Biotechnology, Department of Bioscience, Graduate School of Science and Technology, Shizuoka University, 836 Ohya, Suruga-ku, Shizuoka, 422-8529, Japan. park.enoch@shizuoka.ac.jp.|Laboratory of Biotechnology, Department of Applied Biological Chemistry, Faculty of Agriculture, Shizuoka University, 836 Ohya, Suruga-ku, Shizuoka, 422-8529, Japan. park.enoch@shizuoka.ac.jp.
Enoch Y
EY
Park
D000818:Animals; D012831:Bombyx / Q000235:genetics / Q000378:metabolism*; D048271:Chitosan / Q000737:chemistry / Q000378:metabolism*; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D007814:Larva / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D014162:Transfection / Q000379:methods*
D011994:Recombinant Proteins; D004247:DNA; D048271:Chitosan
BmNPV bacmid; Chitosan; Chitosan–DNA; Nanocomplex; Nanocomplexes; Recombinant protein; Silkworm; β1,3-N-acetylglucosaminyltransferase