Remobilizing deleted piggyBac vector post-integration for transgene stability in silkworm

Area of Science:

• Genetics and Molecular Biology
• Transposable Elements
• Gene Delivery Systems

Background:

• Transgene stability is crucial for the application of transgenic animals.
• The piggyBac system is widely used for gene transfer but can lead to instability.
• Efficient methods for ensuring long-term transgene stability are needed.

Purpose of the Study:

• To construct a modified piggyBac vector for post-integration deletion of transposon sequences.
• To enhance transgene stability in the silkworm (Bombyx mori) genome.
• To evaluate the efficacy of the modified vector in preventing transposon remobilization.

Main Methods:

• Construction of a modified piggyBac vector with specific terminal repeat sequences (L1/R1 and L2/R1).
• Expression of piggyBac transposase (hsp70-PIG) via helper vector or integrated into the modified vector.
• Generation of transgenic silkworms and assessment of transgene integration and stability.

Main Results:

• Shortening L2 sequences improved the transformation frequency of the L1/R1 cassette.
• Post-integration remobilization of the L2/R1 cassette impaired the transposon structure.
• Transgenes linked with impaired transposons remained stable in the genome, even with exogenous transposase presence.

Conclusions:

• The modified piggyBac vector effectively mediates post-integration deletion of transposable elements.
• This strategy guarantees true transgene stabilization in the silkworm genome.
• The developed vector is a valuable tool for transgenic silkworm research and other piggyBac-transformed organisms.