Transgene excision in pollen using a codon optimized serine resolvase CinH-RS2 site-specific recombination system

Area of Science:

• Plant Biotechnology
• Molecular Biology
• Genetics

Background:

• Transgene escape from genetically modified crops poses environmental and regulatory challenges.
• Pollen is a primary vector for transgene flow, necessitating strategies to prevent gene dispersal.

Purpose of the Study:

• To develop and evaluate a novel system for transgene excision from plant pollen.
• To enhance biocontainment of transgenic crops by minimizing transgene escape.

Main Methods:

• Construction of a transgene excision vector with serine recombinase CinH and recognition sites RS2.
• Utilized a pollen-specific LAT52 promoter for controlled expression of CinH recombinase.
• Employed green fluorescent protein (GFP) as a marker for transgene excision, assessed by flow cytometry (FCM).

Main Results:

• The CinH-RS2 recombination system effectively recognized and excised transgenes in pollen.
• Transgenic events with the system showed a significant reduction in GFP-synthesizing pollen (<1%) compared to controls (~70%).
• Flow cytometry analysis confirmed high efficiency of transgene excision from pollen.

Conclusions:

• The CinH-RS2 recombination system is a highly effective tool for transgene biocontainment in crops.
• This approach significantly reduces the risk of transgene escape via pollen.
• The developed system offers a promising solution for environmental safety in transgenic agriculture.