Advancements in Persian walnut tissue culture and genetic engineering: bridging traditional methods with modern...

This review summarizes major advances in Persian walnut biotechnology, emphasizing progress in propagation, somatic embryogenesis, genome editing, and computational tools while outlining key challenges for large-scale propagation and genetic improvement. In vitro culture is fundamental for uniform and large-scale propagation of Persian walnut. Over the past decades, significant improvements have enhanced plant adaptability and survival during transfer to ex vitro environments. Commonly used explants, such as shoot buds, nodal segments, and shoot tips, show variable success depending on genetic, physiological, and environmental factors, as well as culture media composition. Somatic embryogenesis and plant regeneration form the basis for several biotechnological approaches, including haploid production for genomic mapping, mutation analysis, and hybrid development. Recent advances in genome editing, particularly CRISPR/Cas9, have accelerated the creation of cultivars with improved rooting ability, enhanced resistance to biotic stresses, and better tolerance to drought and salinity. Moreover, the integration of machine learning and computational tools has facilitated high-throughput phenotyping, reducing experimental time and cost. Despite these achievements, challenges such as genotype-dependent recalcitrance, oxidative browning, and low transformation efficiency continue to limit large-scale applications. Addressing these obstacles through optimized culture systems and molecular tools will be essential for realizing the full potential of Persian walnut biotechnology. This review provides an integrated overview of recent advances, identifies persistent challenges, and highlights future directions for improving propagation efficiency and accelerating genetic enhancement in this valuable tree species.