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An efficient strategy for producing a stable, replaceable, highly efficient transgene expression system in silkworm,

Dingpei Long1, Weijian Lu1, Yuli Zhang2

  • 1State Key Laboratory of Silkworm Genome Biology, Key Laboratory for Sericulture Functional Genomics and Biotechnology of Agricultural Ministry, Southwest University, Chongqing 400716, People's Republic of China.

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Researchers developed a stable transgene expression system in silkworms (Bombyx mori) by eliminating transposon sequences. This method enables efficient, site-specific gene integration for optimized protein production and potential biocontrol applications.

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Area of Science:

  • Biotechnology
  • Genetics
  • Entomology

Background:

  • Transposon-based systems in silkworms often suffer from random insertion and instability.
  • Efficient and stable transgene expression is crucial for biotechnological applications in insects.

Purpose of the Study:

  • To develop a stable, efficient, and replaceable transgene expression system in silkworms (Bombyx mori).
  • To overcome the limitations of random insertion and post-integration instability associated with transposons.

Main Methods:

  • Developed a strategy combining transposon sequence elimination, site-specific recombination, and optimized fibroin H-chain expression.
  • Generated transgenic silkworm strains, selected TS1-RgG2 for high protein expression, and eliminated transposon sequences via heat-shock-induced transposase expression.
  • Utilized recombinase-mediated cassette exchange (RMCE) with the phiC31/att system for site-specific integration of new genes.

Main Results:

  • Created a stable transgenic silkworm strain (TS3-g2) containing only the desired expression cassette.
  • Achieved up to 16% (w/w) target protein in cocoons from the initial transgenic strain (TS1-RgG2).
  • Demonstrated the potential for integrating other genes of interest into a specific locus using RMCE.

Conclusions:

  • The developed strategy provides a stable and efficient transgene expression system in silkworms.
  • This system facilitates optimized exogenous protein expression and gene function analysis by controlling for position effects.
  • The approach is applicable to other lepidopteran insects, enhancing the ecological safety of transgenic strains for biocontrol.