RNA interference (RNAi) is a conserved mechanism for regulating gene expression in eukaryotes, and RNAi-based strategies have been successfully applied for crop protection against pests and pathogens. These strategies include host-induced gene silencing (HIGS), spray-induced gene silencing (SIGS), and microbe-induced gene silencing (MIGS). However, challenges remain in dsRNA design, large-scale production, and delivery systems. dsRNA design is critical for effective gene silencing, and factors such as RNA structure and target accessibility significantly influence efficiency. The stability and persistence of dsRNA in the environment are also important for effective application. Large-scale production of dsRNA is essential for commercial use, and microbial fermentation and cell-free platforms offer cost-effective solutions. Delivery of dsRNA to target organisms is another challenge, with various nanocarriers and natural systems being explored. Exogenous dsRNAs can induce host immunity, which may affect the efficacy of biopesticides. Additionally, inherited RNAi through transgenerational silencing has been observed in Caenorhabditis elegans, suggesting potential for long-lasting protection against pests and pathogens. Despite these advances, further research is needed to address the challenges and improve the lab-to-field transition of RNAi-based crop protection strategies.RNA interference (RNAi) is a conserved mechanism for regulating gene expression in eukaryotes, and RNAi-based strategies have been successfully applied for crop protection against pests and pathogens. These strategies include host-induced gene silencing (HIGS), spray-induced gene silencing (SIGS), and microbe-induced gene silencing (MIGS). However, challenges remain in dsRNA design, large-scale production, and delivery systems. dsRNA design is critical for effective gene silencing, and factors such as RNA structure and target accessibility significantly influence efficiency. The stability and persistence of dsRNA in the environment are also important for effective application. Large-scale production of dsRNA is essential for commercial use, and microbial fermentation and cell-free platforms offer cost-effective solutions. Delivery of dsRNA to target organisms is another challenge, with various nanocarriers and natural systems being explored. Exogenous dsRNAs can induce host immunity, which may affect the efficacy of biopesticides. Additionally, inherited RNAi through transgenerational silencing has been observed in Caenorhabditis elegans, suggesting potential for long-lasting protection against pests and pathogens. Despite these advances, further research is needed to address the challenges and improve the lab-to-field transition of RNAi-based crop protection strategies.