The Use of CRISPR Technologies for Crop Improvement in Maize

Abstract

AbstractBeing one of the most widely grown cereal species, maize is an agronomically important crop. Traditional plant breeding and the introduction of transgenes have been successfully used to improve maize increasing yield and permitting the development of novel herbicide and insect resistance traits. To keep pace with global food demands and promote sustainability, a new technique for improving crops has emerged, genome editing. Here, researchers have developed tools that cut genomic DNA sequences at defined sites allowing the genetic blueprint of plants to be edited in a targeted fashion. Among these tools, RNA-guided CRISPR-associated (Cas) nucleases have risen to the forefront based on their robustness and ease of re-programmability. In this chapter, we describe how Cas nucleases can be used to introduce genetic variation, either transgenic or non-transgenic, in maize through the creation and repair of chromosomal DNA double-strand breaks (DSBs). Using this approach, we first illustrate how two Cas9-induced DSBs can be used to cleanly excise an entire gene to alter starch composition in maize kernels. Next, we show how transgenes can be inserted at Cas9 DSB sites using homology-directed repair and highlight how this approach can be used to engineer complex trait loci that minimize the risk for endogenous gene disruption and simplify trait evaluation, stacking, and breeding. Next, in lieu of DSBs, we show how CRISPR-Cas enzymes can be engineered to bind chromosomal targets and modulate maize gene expression. For this, the RNA-guided DNA-binding functionality of a new CRISPR system, termed type I cascade, and nuclease-deficient versions of Cas9 were combined with domains that modulate maize gene transcription, altogether, providing new strategies for trait expression. Finally, we discuss how the discovery of new CRISPR-Cas proteins and recruitment of novel domains to nucleic acid targets are leading to the development of new techniques to introduce beneficial changes for maize improvement.KeywordsZea maysWaxy cornMaize transformationComplex trait locidCas9