Agrobacterium-delivered CRISPR/Cas9 for Targeted Mutagenesis in Maize

CRISPR/Cas9 is a powerful genome editing tool in several organisms. Although simpler than other nuclease-based genome editing tools, optimization of CRISPR/Cas9 considers the DNA delivery and the tissue regeneration methods of species to be edited. Si Nian Char of the Iowa State University, together with a team of scientists, reported the ISU Maize CRISPR, which uses Agrobacterium-delivered CRISPR/Cas9 to achieve high-frequency targeted mutagenesis in maize.

The system is composed of an Escherichia coli cloning vector and an Agrobacterium binary vector. It can be used to clone up to four guide RNAs for single or multiple gene targeting. The team evaluated the system for mutagenesis frequency and heritability using four maize genes in two duplicated pairs. T₀ generation transgenic events with any combination of mutation in any of the two loci occurred at rates over 70%.

In the T₁ generation, individuals with only the desired mutant alleles and without the CRISPR/Cas9 transgene could be generated. Double infection of embryos by combining two individual Agrobacterium strains carrying different Cas9/gRNA modules can also be performed to save resources. The ISU Maize CRISPR can be an effective tool for targeted mutagenesis in maize.

For more on this new breeding technology, read the article in Plant Biotechnology Journal.