Transgene-Free Genome Editing for Biotic and Abiotic Stress Resistance in Sugarcane: Prospects and Challenges

Sugarcane (Saccharum spp.) is one of the most valuable food and industrial crops. Its production is constrained due to major biotic (fungi, bacteria, viruses and insect pests) and abiotic (drought, salt, cold/heat, water logging and heavy metals) stresses. The ever-increasing demand for sugar and biofuel and the rise of new pest and disease variants call for the use of innovative technologies to speed up the sugarcane genetic improvement process. Developing new cultivars through conventional breeding techniques requires much time and resources. The advent of CRISPR/Cas genome editing technology enables the creation of new cultivars with improved resistance/tolerance to various biotic and abiotic stresses. The presence of genome editing cassette inside the genome of genome-edited plants hinders commercial exploitation due to regulatory issues. However, this limitation can be overcome by using transgene-free genome editing techniques. Transgene-free genome editing approaches, such as delivery of the RNPs through biolistics or protoplast fusion, virus-induced genome editing (VIGE), transient expression of CRISPR/Cas reagents through Agrobacterium-mediated transformation and other approaches, are discussed. A well-established PCR-based assay and advanced screening systems such as visual marker system and Transgene killer CRISPR system (TKC) rapidly identify transgene-free genome edits. These advancements in CRISPR/Cas technology speed up the creation of genome-edited climate-smart cultivars that combat various biotic and abiotic stresses and produce good yields under ever-changing conditions.

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