CRISPR-enabled functional genomics for bolstering plant tolerance to abiotic and biotic stress; a comprehensive...

Climate change intensifies abiotic stresses including salinity, drought, and extreme temperatures alongside biotic threats such as pathogens and insect pests, collectively undermining global crop productivity and food security. Salinity and drought alone affect 20-50% of irrigated soils, with projections indicating that nearly half of global farmland could become saline by mid-century. Conventional breeding and earlier genome editing tools zinc finger nucleases (ZFNs), and transcription activator-like effector nucleases (ZFNs, TALENs) are constrained by genetic diversity limitations, technical complexity, and slow trait deployment. The CRISPR-Cas9 system has emerged as a transformative platform offering superior precision, efficiency, scalability, and affordability for crop improvement. This review systematically examines how CRISPR-Cas9 enables targeted engineering of stress tolerance in major crops (rice, wheat, maize, tomato, barley) through gene knockout and knock-in strategies. Key applications include editing transcription factors (ART1, DRO1, OsDST) for drought and salinity tolerance, modifying transporter genes (OsHMA2, OsNramp5) for heavy metal detoxification, and disrupting susceptibility genes (MLO, OsERF922, CsLOB1) for broad-spectrum disease and pest resistance. Beyond direct editing, we highlight emerging synergies with functional genomics, multi-omics integration, and high-throughput phenotyping to accelerate target discovery and validation. A central focus is placed on nanobiotechnology-enabled CRISPR delivery systems, including lipid nanoparticles (LNPs), exosomes, and engineered nanocarriers that overcome the plant cell wall barrier a major bottleneck in plant genetic transformation. These platforms enable efficient, genotype-independent delivery of ribonucleoprotein (RNP) complexes, facilitating DNA-free editing for sustainable crop protection. By integrating CRISPR-based precision with advances in nanodelivery and molecular breeding, this review outlines a road-map for developing climate-resilient, high-yielding, and nutritionally enhanced crops to safeguard global agricultural sustainability.