Leaf transformation for efficient random integration and targeted genome modification in maize and sorghum

Area of Science:

• Plant Biotechnology
• Genetics and Genomics
• Agricultural Science

Background:

• Traditional grass transformation methods using immature embryos limit applications to major crops.
• Low success rates in alternative explants, like leaf tissue, hinder broad genome editing adoption in Poaceae.
• Previous leaf transformation using Wuschel2 (Wus2) and Babyboom (Bbm) supported basic mutagenesis but not complex genome editing requiring extensive screening.

Purpose of the Study:

• To enhance leaf transformation efficiency in maize and sorghum using improved Wus2/Bbm expression.
• To enable complex genome editing applications, such as gene dropouts and targeted insertions, in grass species.
• To assess the potential of a maize-optimized Wus2/Bbm construct for broad-spectrum transformation across diverse grass species.

Main Methods:

• Overexpression of Wuschel2 (Wus2) and Babyboom (Bbm) genes in leaf tissues of maize and sorghum.
• Application of Cas9-mediated genome editing for gene dropouts and targeted gene insertion.
• Testing a maize-optimized Wus2/Bbm construct in eight grass species from four subfamilies.

Main Results:

• Enhanced Wus2/Bbm expression significantly improved leaf transformation efficiency in maize and sorghum.
• Successful recovery of plants with Cas9-mediated gene dropouts and targeted gene insertions.
• Production of embryogenic callus and regenerated plantlets across eight species using the optimized Wus2/Bbm construct.

Conclusions:

• Enhanced Wus2/Bbm expression provides a robust method for leaf transformation and genome editing in maize and sorghum.
• The developed Wus2/Bbm system shows promise as a universal transformation and genome editing strategy for the Poaceae family.
• This advancement facilitates complex genome editing applications, accelerating crop improvement in diverse grass species.