Optimized prime editing efficiently generates heritable mutations in maize

Area of Science:

• Plant biotechnology
• Gene editing technologies
• Maize genetics

Background:

• Low efficiency of prime editing (PE) hinders its application in maize.
• Recent advancements in mammalian cells and rice improved PE efficiency through optimized prime-editing guide RNAs (pegRNAs), prime editor (PE) proteins, and cellular factors.

Purpose of the Study:

• To evaluate the efficacy of optimized prime editing systems in maize.
• To overcome the efficiency limitations of prime editing in maize for genetic applications.

Main Methods:

• Testing of optimized prime editor (PE) systems, including the ePE5max system, in maize.
• Utilizing PEmax, engineered pegRNAs (pegRNA-evopreQ.1), nicking single guide RNAs (sgRNAs), and MLH1dn.
• Assessing the generation of heritable mutations conferring herbicide resistance.

Main Results:

• The ePE5max system demonstrated high efficiency in generating heritable mutations in maize.
• Successfully created mutations conferring resistance to EPSPS, ALS, and ACCase inhibiting herbicides.
• Confirmed the system's capability for both homozygous and heterozygous mutations in target genes.

Conclusions:

• The ePE5max system significantly enhances prime editing efficiency in maize.
• This advancement removes the primary obstacle to utilizing prime editing in maize.
• Enables precise genetic modifications for crop improvement and trait development in maize.