Transgenic Research

, Volume 19, Issue 3, pp 473–487

Construction of a binary transgenic gene expression system for recombinant protein production in the middle silk gland of the silkworm Bombyx mori

Authors

  • Ken-ichiro Tatematsu
    • Transgenic Silkworm Research CenterNational Institute of Agrobiological Sciences
  • Isao Kobayashi
    • Transgenic Silkworm Research CenterNational Institute of Agrobiological Sciences
  • Keiro Uchino
    • Transgenic Silkworm Research CenterNational Institute of Agrobiological Sciences
  • Hideki Sezutsu
    • Transgenic Silkworm Research CenterNational Institute of Agrobiological Sciences
  • Tetsuya Iizuka
    • Transgenic Silkworm Research CenterNational Institute of Agrobiological Sciences
  • Naoyuki Yonemura
    • Transgenic Silkworm Research CenterNational Institute of Agrobiological Sciences
    • Transgenic Silkworm Research CenterNational Institute of Agrobiological Sciences
Original Paper

DOI: 10.1007/s11248-009-9328-2

Cite this article as:
Tatematsu, K., Kobayashi, I., Uchino, K. et al. Transgenic Res (2010) 19: 473. doi:10.1007/s11248-009-9328-2

Abstract

To construct an efficient system for the production of recombinant proteins in silkworm (Bombyx mori), we investigated the promoter activity of the silkworm sericin 1, 2, and 3 genes (Ser1, Ser2, and Ser3) using a GAL4/UAS binary gene expression system in transgenic silkworm. The promoter activity of the upstream region of Ser1 was strong, yielding high expression of an enhanced green fluorescent protein (EGFP) transgene in the middle and posterior regions of the middle silk gland (MSG) after day 2 of the fifth instar. The Ser3 upstream region exhibited moderate promoter activity in the anterior MSG, but the Ser2 upstream region did not exhibit any promoter activity. Since the strongest promoter activity was observed for Ser1, we devised a system for the production of recombinant proteins using a GAL4Ser1 promoter construct (Ser1-GAL4). Transgenic silkworms harboring both the Ser1-GAL4 construct and the previously reported upstream activating sequence (UAS)–EGFP construct, which contains the TATA box region of the Drosophilahsp70 gene, yielded approximately 100 μg EGFP per larva. When we then analyzed the TATA box region, signal peptide, and intron sequences for their effects on production from the UAS-EGFP construct, we found that the optimization of these sequences effectively increased production to an average of 500 μg EGFP protein per transgenic larva. We conclude that this binary system is a useful tool for the mass production of recombinant proteins of biomedical and pharmaceutical interest in silkworm.

Keywords

SilkwormBombyxTransgenicBioreactorRecombinant protein

Copyright information

© Springer Science+Business Media B.V. 2009