TILLING and Eco-TILLING: Reverse Genetics Approaches for Crop Improvement

The increasing challenges posed by climate change, resource degradation, and the demand for sustainable agriculture necessitate advanced yet accessible tools for crop improvement. TILLING and its variant, Eco-TILLING, are non-transgenic reverse genetics approaches that enable the identification of induced and naturally occurring genetic variation across diverse crop species. TILLING combines chemical mutagenesis with high-throughput mutation detection to identify functional alleles in specific genes, while Eco-TILLING facilitates the discovery of natural polymorphisms in germplasm collections and wild relatives. These techniques effectively bridge functional genomics and applied plant breeding, particularly in polyploid, orphan, and vegetatively propagated crops. However, limitations include the low frequency of true knockout mutations, the predominance of silent or weak-effect alleles, the need for extensive population screening, and the chimeric nature of M1 plants that complicates early-generation selection. Despite these constraints, TILLING-based approaches remain valuable tools for modern breeding programs, with strong regulatory acceptance and adaptability across agro-ecological conditions.