Genetic Transformation of Recalcitrant Upland Switchgrass Using Morphogenic Genes

Area of Science:

• Plant Biotechnology
• Molecular Biology
• Bioenergy Crops

Background:

• Upland switchgrass cultivars are recalcitrant to genetic transformation, hindering biotechnological applications.
• Efficient genetic modification is crucial for improving switchgrass as a biofuel feedstock.

Purpose of the Study:

• To establish an efficient and reproducible genetic transformation protocol for upland switchgrass cultivars.
• To enable gene function analysis and germplasm enhancement in recalcitrant switchgrass.

Main Methods:

• Ectopic overexpression of morphogenic genes (Baby boom and Wuschel2) in upland switchgrass cultivars.
• Agrobacterium-mediated transformation using auxotrophic strains LBA4404Thy- and EHA105Thy-.
• Selection of transgenic events via GFP expression and confirmation using PCR and herbicide resistance assays.

Main Results:

• Achieved up to 6% transformation efficiency in the 'Summer' cultivar using LBA4404Thy-.
• LBA4404Thy- consistently outperformed EHA105Thy- in transformation frequency.
• Successfully confirmed transgene integration and expression, and demonstrated gene removal via Cre-Lox system.

Conclusions:

• Developed a robust transformation protocol for recalcitrant upland switchgrass.
• This method facilitates genetic studies and biotechnological improvement of switchgrass for bioenergy.
• Opens avenues for enhanced germplasm development in switchgrass.