Seed priming is to expose seeds to specific compounds to enhance seed germination. Few studies of plant immune activation through seed priming have been conducted. Here, we introduce an emerging technology that combines seed priming with elicitation of plant immunity using biologically active compounds. This technology is named 'seed defense biopriming' (SDB). We prepared heat-stable metabolites from 1,825 root-associated Bacillus spp. isolated from the rhizosphere in South Korea. These preparations were tested for their ability to induce SDB in cucumber and pepper seeds and trigger plant immunity. SDB with heat-stable metabolites of the selected Bacillus gaemokensis strain PB69 significantly reduced subsequent bacterial diseases under in vitro and field conditions and increased fruit yield. Transcriptional analysis of induced resistance marker genes confirmed the upregulation of salicylic acid, ethylene, and jasmonic acid signaling. Mortality of the insect pest Spodoptera litura increased when larvae fed on SDB-treated cucumber tissues. Analysis of the causative bacterial metabolites identified a leucine-proline cyclodipeptide and a commercially obtained leucine-proline cyclodipeptide induced similar results as treatment with the bacterial preparation. Our results indicate that SDB treatment with the heat-stable bacterial metabolite effectively elicited immunity and controlled disease in seedlings to whole plants, thereby increasing yield even under field conditions.